Methods and compositions for treating conditions associated with an abnormal inflammatory response

ABSTRACT

This disclosure features chemical entities (e.g., a compound exhibiting activity as a mitochondrial uncoupling agent or a pharmaceutically acceptable salt and/or hydrate and/or cocrystal thereof, e.g., a compound, such as niclosamide or a pharmaceutically acceptable salt and/or hydrate and/or cocrystal thereof, e.g., a compound, such as a niclosamide analog, or a pharmaceutically acceptable salt and/or hydrate and/or cocrystal thereof) that are useful, e.g., for treating one or more symptoms of a pathology characterized by an abnormal inflammatory response (e.g., inflammatory bowel diseases) in a subject (e.g., a human). This disclosure also features compositions as well as other methods of using and making the same.

CROSS REFERENCE TO RELATED APPLICATIONS

This application is a continuation of U.S. application Ser. No.16/173,667, filed Oct. 29, 2018, which is a continuation of U.S.application Ser. No. 15/255,102, filed Sep. 1, 2016, which claims thebenefit of United States Provisional Application No. 62/213,016, filedon Sep. 1, 2015 and U.S. Provisional Application No. 62/241,508, filedon Oct. 14, 2015; each of these prior applications is incorporatedherein by reference in its entirety.

TECHNICAL FIELD

This disclosure features chemical entities (e.g., a compound exhibitingactivity as a mitochondrial uncoupling agent or a pharmaceuticallyacceptable salt, and/or hydrate, and/or cocrystal, and/or drugcombination thereof; e.g., a compound, such as niclosamide or apharmaceutically acceptable salt, and/or hydrate, and/or cocrystal,and/or drug combination thereof; e.g., a compound, such as a niclosamideanalog, or a pharmaceutically acceptable salt, and/or hydrate, and/ordrug combination, and/or cocrystal thereof) that are useful, e.g., fortreating one or more symptoms of a pathology characterized by anabnormal inflammatory response (e.g., inflammatory bowel diseases) in asubject (e.g., a human). This disclosure also features compositions aswell as other methods of using and making the same.

BACKGROUND

Ulcerative colitis (UC) and Crohn's disease (CD) are the predominantchronic, inflammatory bowel diseases (IBD) in humans. These disordersare autoimmune in nature and occur in the absence of infection. IBDeffects up to 2,000,000 Americans (increasing ˜15% annually) and it isassociated with unacceptably high rates of morbidity and mortality. IBDis also a significant burden on the U.S. health care system as the mosteffective treatments are biological drugs that are quite costly.

IBD occurs as the result of inappropriate immune responses ingenetically susceptible individuals mediated by complex interactionsbetween environmental stimuli, microbial factors, and the intestinalimmune system. The hallmark of IBD is represented by excessive immuneresponses that mediate gastrointestinal tissue damage, either directlyor through the release of soluble, pro-inflammatory mediators.

T cells are a type of immune cell that infiltrate the intestinal mucosaand are key drivers of gastrointestinal tissue damage in IBD. Thesecells persist and accumulate in the intestinal mucosa because normalphysiologic mechanisms designed to censor or eliminate activated T cellsare inoperative in the context of IBD. While the exact basis for T cellaccumulation in IBD is not fully elucidated, chronic activation bymicrobial stimuli along with the cytokine milieu at the sites ofinflammation within gastrointestinal tissue are thought to be important.Regardless of how these cells persist, enhancing T cell death in theintestinal mucosa is linked with resolution of IBD and drugs that aremost effective in managing IBD function (in part), by killing pathogenicT cells resident in the gut.

Although different forms of IBD show pathophysiological and clinicaldifferences, the therapeutic approach to managing IBD shares many commonelements. Medical management of IBD is largely empirical, employinganti-inflammatory or immunosuppressive drugs. Salicylazosulfapyridineand 5-aminosalicylic acid are used to treat mild IBD and as maintenancetherapy if disease remission can be achieved. Corticosteroids are usedin patients with moderate to severe disease. However, clinical remissioncan only be obtained in ˜60% of patients, and just about half of thesestay in remission after treatment is discontinued. This last point issignificant because long-term use of corticosteroids carries asignificant risk of serious side effects.

Immunosuppressive drugs can also be used to treat moderate to severecases of IBD, often as a replacement for steroid therapy. However,immunosuppressive drugs (e.g., azathioprine) usually cannot ensurecontrol of symptoms, and treatment is accompanied by numerouscontraindications and severe side effects.

Drugs that often show the best efficacy in treating IBD are systemicallyadministered (via injection or infusion) monoclonal antibodies thatblock TNF-alpha, a pro-inflammatory cytokine overproduced during allforms of IBD (e.g., UC, CD, graft-versus-host disease, celiac disease,iatrogenic colitis such as that induced by checkpoint inhibitors, etc.).Reducing levels of TNF-alpha in the context of IBD has two consequences.First, as an inflammatory cytokine, TNF-alpha mediates tissue damage.Second, high levels of TNF-alpha help disease causing T cells to surviveand blocking TNF-alpha activity eventually leads to T cell death.Indeed, the induction of cell death by anti-TNF-alpha drugs likeinfliximab can predict clinical improvement in patients.

Although effective, use of anti-TNF-alpha drugs is associated withsevere, systemic side effects including, re-activation of latentpathogens, hypersensitivity phenomena, cancer, and the formation ofautoantibodies. Some patients are inherently resistant to anti-TNF-alphadrugs and over time, almost half of all patients that do show aresponse, develop resistance.

From the foregoing it is clear that there is need for new drugs to treatIBD that are more effective, less toxic, less expensive, and moreconvenient to administer versus standard of care.

Niclosamide (5-chloro-N-(2-chloro-4-nitrophenyl)-2-hydrobenzamide) is ahalogenated salicylanilide that belongs to a group of medicines known asanthelmintics. Anthelmintics are medicines used in the treatment of worminfections. Niclosamide, which has low systemic bioavailabilty and anexcellent safety profile, is used to treat broad or fish tapeworm, dwarftapeworm, and beef tapeworm infections. It is believed that Niclosamideinhibits oxidative phosphorylation and stimulates adenosinetriphosphatase activity in the mitochondria of cestodes (e.g.,tapeworm), killing the scolex and proximal segments of the tapeworm bothin vitro and in vivo (see, Li, Y., et al., Cancer Lett. 2014 349,8-14.).

Recent studies have also identified other potential uses of niclosamide;e.g., as a potential anticancer agent (Id.); and as an agent fortreating, preventing and/or alleviating the symptoms of type II diabetesand diabetes-related disorders or complications (see, e.g., WO2012/068274). U.S. Pat. No. 8,148,328 discloses that niclosamideenhances the oral bioavailability of certain peptides.

SUMMARY

This disclosure features chemical entities (e.g., a compound exhibitingactivity as a mitochondrial uncoupling agent or a pharmaceuticallyacceptable salt, and/or hydrate, and/or cocrystal, and/or drugcombination thereof; e.g., a compound, such as niclosamide or apharmaceutically acceptable salt, and/or hydrate, and/or cocrystal,and/or drug combination thereof; e.g., a compound, such as a niclosamideanalog, or a pharmaceutically acceptable salt, and/or hydrate, and/ordrug combination, and/or cocrystal thereof) that are useful, e.g., fortreating one or more symptoms of a pathology characterized by anabnormal inflammatory response (e.g., inflammatory bowel diseases) in asubject (e.g., a human). This disclosure also features compositions aswell as other methods of using and making the same.

This disclosure is based, in part, on the finding that niclosamide killspathogenic T cells isolated from IBD patients and is effective in murinemodels of IBD. While not wishing to be bound by theory, it is believedthat the chemical entities described herein (e.g., niclosamide or apharmaceutically acceptable salt and/or hydrate and/or cocrystalthereof) uncouple mitochondrial respiration from oxidativephosphorylation in one or more T cells, thereby disrupting themitochondrial energy cycle in the one or more T cells and inducing celldeath of the one or more T cells (e.g., activated T cells). It has beensurprisingly found that the chemical entities described hereinselectively target and kill T cells associated with pathologiescharacterized by an abnormal inflammatory response (e.g., pathogenic Tcells in the intestinal mucosa).

The chemical entities, methods, and compositions described herein notonly provide treatment options that are highly efficient and effectiveat killing T cells, but also ones that address the toxicity, cost, andconvenience issues associated with some standard methods of treatment.

In certain embodiments, the methods described herein can be carried outusing niclosamide, a small molecule that has an established and goodsafety profile and is an FDA approved anthelmintic drug.

Additionally, the chemical entities described herein can be readily andefficiently administered locally, such that the resultant systemicbioavailability of the administered chemical entity is relatively low,and the resultant local bioavailability of the administered chemicalentity is relatively high. Local (non-systemic) administration of thechemical entity at a desired area of treatment (e.g., gastrointestinaltract) significantly reduces the likelihood that a patient willexperience systemic toxicities associated with some current standards ofcare. The foregoing can be achieved, for example, by selecting chemicalentities having a relatively low oral bioavailability (F) and/or byemploying formulations that are chemically and/or structurallypredisposed to minimize systemic exposure of the chemical entity (e.g.,the formulations can be designed to release the chemical entity at a pHthat is present in the target area of the GI tract).

In view of the foregoing advantages and features delineated above, thechemical entities, methods, and compositions described herein are alsoexpected to be functional in diverse patient populations and/or lesssensitive to blocks in cell death mechanisms. Further, the ability toutilize traditional small molecules, such as niclosamide, can helpreduce cost and facilitate patient administration.

In some embodiments, the methods and compositions described herein aresuitable for use in combination therapy with various other therapeuticregimens (e.g., chemotherapy and/or radiation). In certain embodiments,the chemical entities and methods described herein can be used to treatside effects produced by such therapeutic regimens, e.g., inflammatorybowel diseases induced by chemotherapeutic immunomodulators, e.g.,checkpoint inhibitors, which in some cases can be prohibitively severe.Additionally, the chemical entities, methods, and compositions describedherein are also expected to be useful in certain treatment-resistantpatient populations, e.g., one that is nonresponsive or resistant totreatment an anti-TNFalpha therapy (e.g., Humira, Enbrel, Remicade).

In one aspect, methods for inducing cell death of one or more T cells(e.g., in the digestive and/or gastrointestinal tract (GI), skin, eyes,or joints), of a subject are provided. The methods include contactingthe one or more T cells with an effective amount of a chemical entity(e.g., a compound exhibiting activity as a mitochondrial uncouplingagent or a pharmaceutically acceptable salt and/or hydrate and/orcocrystal thereof; e.g., a compound, such as niclosamide or apharmaceutically acceptable salt and/or hydrate and/or cocrystalthereof; e.g., a compound, such as a niclosamide analog, or apharmaceutically acceptable salt and/or hydrate and/or cocrystalthereof) as defined anywhere herein.

In another aspect, methods for treating a subject having a conditionassociated with unregulated (abnormal, elevated) recruitment and/orretention of one or more T cells (e.g., at the digestive and/orgastrointestinal tract (GI), skin, eyes, or joints) of the subject areprovided. The methods include contacting the one or more T cells with aneffective amount of a chemical entity (e.g., a compound exhibitingactivity as a mitochondrial uncoupling agent or a pharmaceuticallyacceptable salt and/or hydrate and/or cocrystal thereof; e.g., acompound, such as niclosamide or a pharmaceutically acceptable saltand/or hydrate and/or cocrystal thereof; e.g., a compound, such as aniclosamide analog, or a pharmaceutically acceptable salt and/or hydrateand/or cocrystal thereof) as defined anywhere herein.

In a further aspect, methods for treating a subject having a conditionassociated with unregulated (abnormal, elevated) activation of one ormore T cells (e.g., in the digestive and/or gastrointestinal tract (GI),skin, eyes, or joints) of the subject are provided. The methods includecontacting the one or more activated T cells with an effective amount ofa cocrystal comprising (i) a mitochondrial uncoupling agent or apharmaceutically acceptable salt and/or hydrate thereof; and (ii) one ormore pharmaceutically acceptable coformers as defined anywhere herein.

In aspect, methods for treating a condition (or one or more symptomsthereof) characterized by an abnormal inflammatory response in a subjectin need thereof are provided (e.g., an autoimmune disorder, e.g., aninflammatory bowel disease). The methods include administering to thesubject an effective amount of a chemical entity (e.g., a compoundexhibiting activity as a mitochondrial uncoupling agent or apharmaceutically acceptable salt and/or hydrate and/or cocrystalthereof; e.g., a compound, such as niclosamide or a pharmaceuticallyacceptable salt and/or hydrate and/or cocrystal thereof; e.g., acompound, such as a niclosamide analog, or a pharmaceutically acceptablesalt and/or hydrate and/or cocrystal thereof) as defined anywhereherein.

In another aspect, methods for treating a condition (or one or moresymptoms thereof) characterized by an abnormal inflammatory response ina subject in need thereof are provided (e.g., an autoimmune disorder,e.g., an inflammatory bowel disease). The methods include topically andlocally administering to the subject an effective amount of a chemicalentity (e.g., a compound exhibiting activity as a mitochondrialuncoupling agent or a pharmaceutically acceptable salt and/or hydrateand/or cocrystal thereof; e.g., a compound, such as niclosamide or apharmaceutically acceptable salt and/or hydrate and/or cocrystalthereof; e.g., a compound, such as a niclosamide analog, or apharmaceutically acceptable salt and/or hydrate and/or cocrystalthereof) as defined anywhere herein.

In a further aspect, methods for treating autoimmune colitis (or one ormore symptoms thereof) in a subject are provided. The methods includetopically and locally administering to the subject an effective amountof a chemical entity (e.g., a compound exhibiting activity as amitochondrial uncoupling agent or a pharmaceutically acceptable saltand/or hydrate and/or cocrystal thereof; e.g., a compound, such asniclosamide or a pharmaceutically acceptable salt and/or hydrate and/orcocrystal thereof; e.g., a compound, such as a niclosamide analog, or apharmaceutically acceptable salt and/or hydrate and/or cocrystalthereof) as defined anywhere herein.

In one aspect, methods for treating a condition (or one or more symptomsthereof) selected from the group consisting of celiac disease, irritablebowel syndrome, mucositis, uveitis, collagenous colitis, lymphocyticcolitis, microscopic colitis, radiation enteritis, rheumatoid arthritis,lupus, scleroderma, psoriasis, cutaneous T-cell lymphoma, acute graftvs. host disease and chronic graft vs. host disease in a subject areprovided. The methods include topically and locally administering to thesubject an effective amount of a chemical entity (e.g., a compoundexhibiting activity as a mitochondrial uncoupling agent or apharmaceutically acceptable salt and/or hydrate and/or cocrystalthereof; e.g., a compound, such as niclosamide or a pharmaceuticallyacceptable salt and/or hydrate and/or cocrystal thereof; e.g., acompound, such as a niclosamide analog, or a pharmaceutically acceptablesalt and/or hydrate and/or cocrystal thereof) as defined anywhereherein.

In one aspect, a cocrystal is provided, which includes: (i) amitochondrial uncoupling agent or a pharmaceutically acceptable saltand/or hydrate thereof, and (ii) one or more pharmaceutically acceptablecoformers.

Definitions

To facilitate understanding of the disclosure set forth herein, a numberof terms are defined below. Generally, the nomenclature used herein andthe laboratory procedures in organic chemistry, medicinal chemistry, andpharmacology described herein are those well-known and commonly employedin the art. Unless defined otherwise, all technical and scientific termsused herein generally have the same meaning as commonly understood byone of ordinary skill in the art to which this disclosure belongs. Eachof the patents, applications, published applications, and otherpublications that are mentioned throughout the specification and theattached appendices are incorporated herein by reference in theirentireties.

The term “digestive tract” is understood to include the mouth, pharynx,esophagus, stomach, small intestine (duodenum, jejunum, ileum), largeintestine (cecum, colon, rectum) and anus.

The term “oral cavity” is understood to include the mouth, the pharynxand the esophagus.

The term “gastrointestinal tract”, or “GI tract” is understood toinclude the stomach, small intestine (duodenum, jejunum, ileum), largeintestine (cecum, colon, rectum) and anus.

The term “acceptable” with respect to a formulation, composition oringredient, as used herein, means having no persistent detrimentaleffect on the general health of the subject being treated.

“API” refers to an active pharmaceutical ingredient.

The terms “effective amount” or “therapeutically effective amount,” asused herein, refer to a sufficient amount of a chemical entity (e.g., acompound exhibiting activity as a mitochondrial uncoupling agent or apharmaceutically acceptable salt and/or hydrate and/or cocrystalthereof; e.g., a compound, such as niclosamide or a pharmaceuticallyacceptable salt and/or hydrate and/or cocrystal thereof; e.g., acompound, such as a niclosamide analog, or a pharmaceutically acceptablesalt and/or hydrate and/or cocrystal thereof) being administered whichwill relieve to some extent one or more of the symptoms of the diseaseor condition being treated. The result includes reduction and/oralleviation of the signs, symptoms, or causes of a disease, or any otherdesired alteration of a biological system. For example, an “effectiveamount” for therapeutic uses is the amount of the composition comprisinga compound as disclosed herein required to provide a clinicallysignificant decrease in disease symptoms. An appropriate “effective”amount in any individual case is determined using any suitabletechnique, such as a dose escalation study.

The term “excipient” or “pharmaceutically acceptable excipient” means apharmaceutically-acceptable material, composition, or vehicle, such as aliquid or solid filler, diluent, carrier, solvent, or encapsulatingmaterial. In one embodiment, each component is “pharmaceuticallyacceptable” in the sense of being compatible with the other ingredientsof a pharmaceutical formulation, and suitable for use in contact withthe tissue or organ of humans and animals without excessive toxicity,irritation, allergic response, immunogenicity, or other problems orcomplications, commensurate with a reasonable benefit/risk ratio. See,e.g., Remington: The Science and Practice of Pharmacy, 21st ed.;Lippincott Williams & Wilkins: Philadelphia, Pa., 2005; Handbook ofPharmaceutical Excipients, 6th ed.; Rowe et al., Eds.; ThePharmaceutical Press and the American Pharmaceutical Association: 2009;Handbook of Pharmaceutical Additives, 3rd ed.; Ash and Ash Eds.; GowerPublishing Company: 2007; Pharmaceutical Preformulation and Formulation,2nd ed.; Gibson Ed.; CRC Press LLC: Boca Raton, Fla., 2009.

The term “pharmaceutically acceptable salt” refers to a formulation of acompound that does not cause significant irritation to an organism towhich it is administered and does not abrogate the biological activityand properties of the compound. In certain instances, pharmaceuticallyacceptable salts are obtained by reacting a compound described herein,with acids such as hydrochloric acid, hydrobromic acid, sulfuric acid,nitric acid, phosphoric acid, methanesulfonic acid, ethanesulfonic acid,p-toluenesulfonic acid, salicylic acid and the like. In some instances,pharmaceutically acceptable salts are obtained by reacting a compoundhaving acidic group described herein with a base to form a salt such asan ammonium salt, an alkali metal salt, such as a sodium or a potassiumsalt, an alkaline earth metal salt, such as a calcium or a magnesiumsalt, a salt of organic bases such as dicyclohexylamine,N-methyl-D-glucamine, tris(hydroxymethyl)methylamine, and salts withamino acids such as arginine, lysine, and the like, or by other methodspreviously determined. The pharmacologically acceptable salt s notspecifically limited as far as it can be used in medicaments. Examplesof a salt that the compounds described hereinform with a base includethe following: salts thereof with inorganic bases such as sodium,potassium, magnesium, calcium, and aluminum; salts thereof with organicbases such as methylamine, ethylamine and ethanolamine; salts thereofwith basic amino acids such as lysine and ornithine; and ammonium salt.The salts may be acid addition salts, which are specifically exemplifiedby acid addition salts with the following: mineral acids such ashydrochloric acid, hydrobromic acid, hydroiodic acid, sulfuric acid,nitric acid, and phosphoric acid:organic acids such as formic acid,acetic acid, propionic acid, oxalic acid, malonic acid, succinic acid,fumaric acid, maleic acid, lactic acid, malic acid, tartaric acid,citric acid, methanesulfonic acid, and ethanesulfonic acid; acidic aminoacids such as aspartic acid and glutamic acid.

The term “pharmaceutical composition” refers to a mixture of a compounddescribed herein with other chemical components (referred tocollectively herein as “excipients”), such as carriers, stabilizers,diluents, dispersing agents, suspending agents, and/or thickeningagents. The pharmaceutical composition facilitates administration of thecompound to an organism. Multiple techniques of administering a compoundexist in the art including, but not limited to: rectal, oral,intravenous, aerosol, parenteral, ophthalmic, pulmonary, and topicaladministration.

The term “subject” refers to an animal, including, but not limited to, aprimate (e.g., human), monkey, cow, pig, sheep, goat, horse, dog, cat,rabbit, rat, or mouse. The terms “subject” and “patient” are usedinterchangeably herein in reference, for example, to a mammaliansubject, such as a human.

The terms “treat,” “treating,” and “treatment,” in the context oftreating a disease or disorder, are meant to include alleviating orabrogating a disorder, disease, or condition, or one or more of thesymptoms associated with the disorder, disease, or condition; or toslowing the progression, spread or worsening of a disease, disorder orcondition or of one or more symptoms thereof. Often, the beneficialeffects that a subject derives from a therapeutic agent do not result ina complete cure of the disease, disorder or condition.

As used herein, the terms “alkyl” and the prefix “alk-” are inclusive ofboth straight chain and branched chain groups and of cyclic groups,i.e., cycloalkyl. A “C₂₋₁₀ alkenyl” refers to a branched or unbranchedhydrocarbon group containing one or more double bonds and having from 2to 10 carbon atoms. A “C₂₋₁₀ alkynyl” refers to a branched or unbranchedhydrocarbon group containing one or more triple bonds and having from 2to 10 carbon atoms. A “C₂₋₆ heterocyclyl” refers to a stable 5- to7-membered monocyclic or 7- to 14-membered bicyclic heterocyclic ringthat is saturated, partially unsaturated or unsaturated (aromatic), andthat consists of 2 to 6 carbon atoms and 1, 2, 3 or 4 heteroatomsindependently selected from the group consisting of N, O, and S andincluding any bicyclic group in which any of the above-definedheterocyclic rings is fused to a benzene ring. A “C₆₋₁₂ aryl” refers toan aromatic group having a ring system comprised of carbon atoms withconjugated electrons (e.g., phenyl). A “C₇₋₁₄ alkaryl” refers to analkyl substituted by an aryl group (e.g., benzyl, phenethyl, or3,4-dichlorophenethyl) having from 7 to 14 carbon atoms A “C₃₋₁₀alkheterocyclyl” refers to an alkyl substituted heterocyclic group. A“C₁₋₁₀ heteroalkyl” refers to a branched or unbranched alkyl, alkenyl,or alkynyl group having from 1 to 10 carbon atoms in addition to one ormore heteroatoms, where one or more methylenes (CH₂) or methines (CH)are replaced by nitrogen, oxygen, sulfur, carbonyl, thiocarbonyl,phosphoryl, or sulfonyl. The term “acyl” refers to a chemical moietywith the formula R—C(O)—, where R is selected from C₁₋₁₀ alkyl, C₁₋₁₀alkenyl, C₁₋₁₀ alkynyl, C₂₋₆ heterocyclyl, C₆₋₁₂ aryl, C₇₋₁₄ alkaryl,C₃₋₁₀ alkheterocyclyl, C₂₋₆ heteroalkyl, and the like. In certainembodiments, acyl is a chemical moiety with the formula R—C(O)—, where Ris selected from C₁₋₁₀ alkyl, C₁₋₁₀ alkenyl, C₁₋₁₀ alkynyl, C₂₋₆heterocyclyl, C₆₋₁₂ aryl, C₇₋₁₄ alkaryl, C₃₋₁₀ alkheterocyclyl, andC₁₋₁₀ heteroalkyl. Each of the foregoing groups can be independentlysubstituted or unsubstituted. Illustrative substituents include alkoxy,aryloxy, sulflhydryl, alkylthio, arylthio, halide, hydroxyl,fluoroalkyl, perfluoralkyl, amino, aminoalkyl, disubstituted amino,quaternary amino, hydroxyalkyl, carboxyalkyl, and carboxyl groups.

The details of one or more embodiments of the invention are set forth inthe accompanying drawings and the description below. Other features andadvantages of the invention will be apparent from the description anddrawings, and from the claims.

DESCRIPTION OF DRAWINGS

FIG. 1 contains graphs showing that Niclosamide induces cell death inlamina propria T cell from active IBD. LPMC (lamina propria mononuclearcells) from IBD subjects were isolated from macroscopically inflamedintestinal area and treated with DMSO or niclosamide (10 μM) for 16hours. Cell death in lamina propria T cell (CD3+) was determined bymeasuring 7-AAD staining by flow cytometry.

FIG. 2 includes graphs and images showing that niclosamide exhibitsrobust efficacy in murine TNBS model of ulcerative colitis whenadministered rectally (locally), but not by intraperitoneal injection(systemically).

FIGS. 3A-3C show the components of a representative enema deliverydevice (FIG. 3A shows the bottle, FIG. 3B shows the breakable capsule,and FIG. 3C shows the rectal cannula (upper arrow) and single flow pack(lower arrow).

FIG. 4A is a graph showing that niclosamide suspension administeredrectally at a dose of 30 mg/kg on days 1 and 2 results in recovery ofbody weight initially last due to TNBS-induced colitis. There is norecovery of weight in untreated or vehicle control treated mice.

FIG. 4B is a graph showing that niclosamide suspension administeredrectally at a dose of 30 mg/kg on days 1 and 2 results in asignificantly lower colitis score compared to vehicle control treatedmice or mice that received TNBS and no other treatment, based on H&Eanalysis of colon biopsies.

FIG. 4C includes graphs that demonstrate expression of inflammatorycytokines in intestinal biopsied tissue detected by real-time PCR. TNBSexposure in presence of vehicle increases expression of TNFa, IFNy andIL-17A compared to EtOH control animals that receive no TNBS.Niclosamide administered rectally at 0.03, 3.0 and 30 mg per kg bodyweight dose-dependently reduces the level of RNA of each cytokinerelative to expression of RNA for β-actin, used as a housekeeping genefor normalization.

FIG. 5 is a graph showing that niclosamide at 5 μM causes a decrease inhuman LPMCs T cells that produce pro-inflammatory cytokines includingTNF, IFN, and IL-17A relative to vehicle only negative control

FIG. 6 is a graph showing that niclosamide at 5 μM causes a decrease inΔΨm in human LPMCs T cells relative to negative control.

DETAILED DESCRIPTION

This disclosure features chemical entities (e.g., a compound exhibitingactivity as a mitochondrial uncoupling agent or a pharmaceuticallyacceptable salt, and/or hydrate, and/or cocrystal, and/or drugcombination thereof; e.g., a compound, such as niclosamide or apharmaceutically acceptable salt, and/or hydrate, and/or cocrystal,and/or drug combination thereof; e.g., a compound, such as a niclosamideanalog, or a pharmaceutically acceptable salt, and/or hydrate, and/ordrug combination, and/or cocrystal thereof) that are useful, e.g., fortreating one or more symptoms of a pathology characterized by anabnormal inflammatory response (e.g., inflammatory bowel diseases) in asubject (e.g., a human). This disclosure also features compositions aswell as other methods of using and making the same.

Chemical Entities

Evaluating Chemical Entities for Activity as Mitochondrial UncouplingAgents

While not wishing to be bound by theory, it is believed that thechemical entities described herein are capable of uncouplingmitochondrial respiration from oxidative phosphorylation in one or moreT cells, thereby disrupting the mitochondrial energy cycle in the one ormore T cells, and inducing cell death of the one or more T cells (e.g.,activated T cells). The ability of a chemical entity to uncouplemitochondrial respiration from oxidative phosphorylation in one or moreT cells can be evaluated using conventional assays that are known in theart.

By way of example, the Jurkat T cell model can be used to study thepotential effects of compounds on T cells in vitro. This cell lineallows investigation of stimuli and mechanisms that regulate T cellmitochondrial function and survival. As T cells, Jurkats have alymphocyte appearance and replicate in culture in suspension. Jurkatsalso contain respiring mitochondria and, as such, response tomitochondrial uncouplers, e.g., niclosamide, may be assessed. Uncouplingis identified and quantified by a detecting a drop in theelectrochemical gradient across the mitochondrial inner membrane (ΔΨm)that is not associated with a corresponding increase in oxidativephosphorylation. Experiments to detect changes in ΔΨm were performed byincluding conditions in which a concentration of oligomycin was added toirreversibly inhibit the F₁F₀-ATPase and block oxidative phosphorylationto demonstrate that the fall in ΔΨm represents uncoupling since itoccurred independent of an increase in mitochondrial oxidativephosphorylation. See Example 1.

As another example, lamina propria mononuclear cells (LPMC) in the humanintestine are comprised in part by T cells which mediate physiologicaland pathological processes including inflammatory bowel disease. LPMCscan be isolated from human tissue biopsies. After isolation LPMCs Tcells remain viable ex vivo under appropriate culture conditions forperiods of time that allow ex vivo experiments. These cells can be usedto investigate mechanisms that regulate their mitochondrial function andsurvival. They contain respiring mitochondria and as such their responseto mitochondrial uncouplers such as niclosamide may be assessed. Thiscellular model can be used in conjunction with oligomycin that blocksoxidative phosphorylation and TMRM to monitor ΔΨm as described inExample 1. See Example 2.

Chemical entities that exhibit mitochondrial uncoupling agent activitycan also include those that exhibit mild uncoupling, which refers to alevel of proton leak that is compensated for by increased mitochondrialoxygen consumption so as to prevent a significant drop in thetransmembrane potential.

Physicochemical Properties of Chemical Entities

In some embodiments, it is advantageous when the resultant systemicbioavailability of the administered chemical entity is relatively low,and the resultant local bioavailability of the administered chemicalentity is relatively high. The foregoing can be achieved, for example,by selecting chemical entities having a relatively low oralbioavailability (F), wherein:F=Fa×Fg×Fhin which Fa=fraction absorbed; Fg=fraction escaping gut metabolism; andFh=fraction escaping hepatic metabolism (see Filipski, K. J., et al.,Current Topics in Medicinal Chemistry, 2013, 13, 776-802). As theskilled artisan will appreciate, the degree of oral bioavailability canbe influenced by various physicochemical attributes, such as molecularweight (“MW”), log P, number of hydrogen bond donors (“HBD”), number ofhydrogen bond acceptors (“HBA”), number of rotatable bonds (“RB”), andpolar surface area (“PSA”). It has been recognized that good oralbioavailability is typically observed in compounds having the followingattributes: MW≤500, Log P≤5, HBD≤5, HBA≤10, rotatable bonds (RB)≤10,PSA≤140 (Id.). Accordingly, a non-limiting strategy for designing andselecting chemical entities having a relatively low oral bioavailability(F) can include selecting physicochemical attributes that conferproperties outside of the preferred oral drug space (Id.).

In some embodiments, the chemical entities described herein (includingtheir pharmaceutically acceptable salts and/or hydrates and/orcocrystals thereof) have an oral bioavailability (F) of less than about50%, or less than about 40%, or less than about 30%, or less than about20%, or less than about 10%, or less than about 5%, or less than about2%, or less than about 1%. In certain embodiments, the chemical entitiesdescribed herein have an oral bioavailability (F) of less than about20%, e.g., less than about 19%, less than about 18%, less than about17%, less than about 16%, less than about 15%, less than about 14%, lessthan about 13%, less than about 12%, less than about 11%, less thanabout 10%, less than about 9%, less than about 8%, less than about 7%,less than about 6%, less than about 5%, less than about 4%, less thanabout 3%, less than about 2%, less than about 1%, or less than about0.5%.

In some embodiments, the chemical entities described herein (includingtheir pharmaceutically acceptable salts and/or hydrates and/orcocrystals thereof) have a relatively low aqueous solubility. Lowaqueous solubility refers to a compound having a solubility in waterwhich is less than or equal to 10 mg/mL, when measured at 20° C. Incertain embodiments, the chemical entities described herein have aqueoussolubility of less than or equal to 900, 800, 700, 600, 500, 400, 300,200 150 100, 90, 80, 70, 60, 50, 40, 30, 20 micrograms/mL, or further10, 5 or 1 micrograms/mL, or further 900, 800, 700, 600, 500, 400, 300,200 150, 100 90, 80, 70, 60, 50, 40, 30, 20, or 10 ng/mL, or less than10 ng/mL when measured at 20° C.

In some embodiments, the chemical entities described herein (includingtheir pharmaceutically acceptable salts and/or hydrates and/orcocrystals thereof) have a relatively low drug permeability.Permeability measurements are based indirectly on the extent ofabsorption of a drug substance in humans and directly on the measurementof rates of mass transfer across human intestinal membrane.Alternatively, non-human systems capable of predicting drug absorptionin humans can be used (such as in-vitro culture methods). A drugsubstance is considered highly permeable when the extent of absorptionin humans is determined to be about 90% or more of the administered dosebased on a mass-balance determination or in comparison to an intravenousdose.

Otherwise, the drug substance is considered to be poorly permeable (see,e.g.,https://books.google.com/books?id=4cfzT2ZY8hUC&pg=PA102&1pg=PA102&dq=low+permeability+drug+definition&source=bl&ots=WXEDT3C0sL&sig=g1laf7e47KJ-SSV4loN8RSs_sM&hl=en&sa=X&ved=0CFAQ6AEwBmoVChMIrv_6oLFxwIVxBmSCh02ugoi#v=onepage&q=low%20permeability%20drug%20definition&f=false).

In some embodiments, the chemical entities described herein can be a BCSclass II drug, or pharmaceutically acceptable salt and/or hydrate and/orcocrystal thereof. In other embodiments, the chemical entities describedherein can be a BCS class IV drug, or pharmaceutically acceptable saltand/or hydrate and/or cocrystal thereof.

Chemical Entities

Niclosamide and Niclosamide Analogs

In some embodiments, the chemical entity can be niclosamide or apharmaceutically acceptable salt and/or hydrate thereof; e.g., acompound, such as a niclosamide analog, or a pharmaceutically acceptablesalt and/or hydrate. Niclosamide analogues refer to compounds, in whichone or more atoms, functional groups, or substructures in niclosamideis/are replaced with one or more different atoms, groups, orsubstructures.

In certain embodiments, the chemical entity can be a compound havingformula I:

where X is N or CR¹⁰; Y is N or CR¹¹; Z is N or CR¹²; and each of R¹,R², R⁵, R⁶, R⁷, R⁸, R⁹, R¹⁰, R¹¹ and R¹² is independently selected fromH, halide (F, Cl, Br, or I). NO₂, OH, OR¹³, SR¹⁴, NR¹⁵R¹⁶, CN, CF₃,C₁₋₁₀ alkyl, C₂₋₁₀ alkenyl, C₂₋₁₀ alkynyl, C₂₋₆ heterocyclyl, C₆₋₁₂aryl, C₇₋₁₄ alkaryl, C₃₋₁₀ alkheterocyclyl, C₁₋₁₀ heteroalkyl, or isdescribed by one of the following formulas:

In compounds of formula I, R³ and R⁴ are independently selected from thegroup consisting of C═O, C═S, C═NR⁴², NH, NR⁴³, CHOR⁴⁴, CH₂, and thelike. Groups R² and R⁴; X and R⁴; R⁵ and R³; R⁹ and R³ may combine toform a six-membered ring, using connections described by one of thegroups:

For compounds of formula I, each E¹ is independently O, S, or NR⁴², eachE² is independently CR⁴⁹R⁵⁰, O or S; each E³ is independently CR⁵¹R⁵²,O, S, or NR⁵³; each Q is, independently, O, S, or NR⁵⁴. R¹³ and R¹⁴ areeach independently, acyl, C₁₋₁₀ alkyl, C₂₋₁₀ alkenyl, C₂₋₁₀ alkynyl,C₂₋₆ heterocyclyl, C₆₋₁₂ aryl, C₇₋₁₄ alkaryl, C₃₋₁₀ alkheterocyclyl,C₂₋₁₀ heteroalkyl; R¹⁸, R²³, R²⁸, R²⁹, R³⁰, R⁴², R⁵⁴ are each,independently, C₁₋₁₀ alkyl, C₂₋₁₀ alkenyl, C₂₋₁₀ alkynyl, C₂₋₆heterocyclyl, C₆₋₁₂ aryl, C₇₋₁₄ alkaryl, C₃₋₁₀ alkheterocyclyl, C₁₋₁₀heteroalkyl; R¹⁵, R¹⁶, R¹⁷, R¹⁹, R²⁰, R²¹, R²², R²⁴, R²⁵, R²⁶, R²⁷, R⁴³,R⁴⁴, R⁴⁵, R⁴⁶, R⁴⁷, R⁴⁸, R⁵¹, R⁵², and R⁵³ are each, independently, H,C₁₋₁₀ alkyl, C₂₋₁₀ alkenyl, C₂₋₁₀ alkynyl, C₂₋₆ heterocyclyl, C₆₋₁₂aryl, C₆₋₁₂, alkaryl, C₁₋₁₀ alkheterocyclyl, C₁₋₁₀ heteroalkyl; R³¹,R³², R³³, R³⁴, R³⁵, R³⁶, R³⁷, R³⁸, R³⁹, R⁴⁰, R⁴¹, R⁴⁹, and R⁵⁰ are each,independently, H-L halide, NO₂, CN, CF₃, C₁₋₁₀ alkyl, C₂₋₁₀ alkenyl,C₂₋₁₀ alkynyl, C₂₋₆ heterocyclyl, C₆₋₁₂ aryl, C₇₋₁₄ alkaryl, C₃₋₁₀alkheterocyclyl, or C₁₋₁₀ heteroalkyl.

In certain embodiments, the chemical entity can be a compound having anyone of formulas XVIII-XXI:

wherein X, Y, Z, E¹, R¹, R⁵, R⁶, R⁷, R⁸, R⁹, R⁴⁷, and R⁴⁸ are as definedabove.

In certain embodiments, the chemical entity can be a compound havingFormula XXII:

wherein R¹, R², R⁵, R⁶, R⁷, R⁸, R⁹, R¹⁰, R¹¹ and R¹² are independentlyselected from the group consisting of H, halide, NO₂, CF₃, OH, acyl, CN,C₁-C₁₀ alkyl (preferably C₁-C₃ alkyl), C₁-C₁₀ heteroalkyl (preferablyC₁-C₃ heteroalkyl), and wherein R³ and R⁴ are as defined above. Incertain embodiments, R³ is C═O, while R⁴ is NH or R³ is NH while R⁴ isC═O. In these and certain other embodiments, only two of R¹, R², R¹⁰,R¹¹, and R¹² are present, and one is H or OH, while the other is halogen(e.g., Cl, Br, or F). In other embodiments, one of R¹, R², R¹⁰, R¹¹, andR¹² is H or OH, one of R¹, R², R¹¹, R¹², and R¹² is halogen (e.g., Cl,Br, or F), and the others are hydrogen.

In these and certain other embodiments, only two of R⁵, R, R⁷, R⁸, andR⁹ are present and these are NO₂ and halogen (e.g., Cl, Br, or F) Inother embodiments, one of R⁵, R⁶, R⁷, R⁸, and R⁹ is NO₂, one of R⁵, R⁶,R⁷, R⁸, and R⁹ is halogen (e.g., Cl, Br, or F), and the rest arehydrogen. In certain embodiments niclosamide analogues include, but arenot limited to niclosamide analogues in which one halogen group isrelocated within the same ring or both halogen groups are relocatedwithin the same ring, niclosamides in which the nitro group is relocatedwithin the same ring, niclosamide analogues where the hydroxyl group isrelocated within the same ring, niclosamide analogues where both halogenand hydroxy and/or nitro groups are relocated while keeping thesubstituents within the aromatic ring, compounds, except having except(3-chloro-4-nitrophenyl) in place of (2-chloro-4-nitrophenyl),niclosamide analogues having a nitro- and a hydroxyl group relocation,niclosamide analogues comprising a single halogen exchange, niclosamideanalogues comprising a double halogen exchange, niclosamide analogscomprising an exchange of Cl— to Br—, niclosamide analogs comprising anexchange of Cl— to F—, and the like.

In certain embodiments the niclosamide analogues include, but are notlimited to compounds according to Formula XXIII:

wherein R¹, R², R³, R⁴, and R⁵, are independently present or absent, andwhen present are independently selected from the group consisting of Cl,Br, alkyl, methyl, hydroxyalkyl, and the like. These analogues are meantto be illustrative and not limiting.

In certain embodiments, the chemical entity can be a compound havingformula XXIV, or a pharmaceutically acceptable salt and/or hydratethereof:

wherein R¹, R², R⁵, R⁶, R⁷, R⁸, R⁹, R¹⁰, R¹¹ and R¹² are independentlyselected from the group consisting of H, halide, NO₂, CF₃, OH, acyl, CN,C₁-C₁₀ alkyl (preferably C₁-C₃ alkyl), and C₁-C₁₀ heteroalkyl(preferably C₁-C₃ heteroalkyl); and wherein R³ is C═O, and R⁴ is NH; orR³ is NH, and R⁴ is C═O, wherein at least one of R¹, R², R⁵, R⁶, R⁷, R⁸,R⁹, R¹⁰, R¹¹ and R¹² is other than H.

In certain of these embodiments, two of R¹, R², R¹⁰, R¹¹ and R¹² areindependently selected from halide, NO₂, CF₃, OH, acyl, CN, C₁-C₁₀ alkyl(preferably C₁-C₃ alkyl), and C₁-C₁₀ heteroalkyl (preferably C₁-C₃heteroalkyl), and the others are H; and two of R⁵, R⁶, R⁷, R⁸, and R⁹are independently selected from halide, NO₂, CF₃, OH, acyl, CN, C₁-C₁₀alkyl (preferably C₁-C₃ alkyl), and C₁-C₁₀ heteroalkyl (preferably C₁-C₃heteroalkyl), and the others are H.

In certain embodiments, the chemical entity can be a compound havingformula XXV, or a pharmaceutically acceptable salt and/or hydratethereof.

wherein one or more of R¹, R², R³, R⁴, and R⁵ is halide, NO₂, CF₃, OH,acyl, CN, C₁-C₁₀ alkyl (preferably C₁-C₃ alkyl), or C₁-C₁₀ heteroalkyl(preferably C₁-C₃ heteroalkyl); and the others are hydrogen.

Examples of niclosamide analogues include, but are not limited to thosedelineated in Tables 1-3.

TABLE 1

TABLE 2

TABLE 3

n = 0; R = Cl n = 1; R = H n = 2; R = H

In certain embodiments, the chemical entity can be a compound havingformula (XXVI):

R¹ represents C₁₋₆alkyl, C₂₋₆alkenyl, C₂₋₆alkynyl, C₃₋₈cycloalkyl,C₄₋₈cycloalkenyl or aryl, all of which may optionally be furthersubstituted with C₁₋₈alkyl, C₃₋₈cycloalkyl, C₄₋₈cycloalkenyl or phenyl;or R¹ represents a bicyclo-C₄₋₁₀alkyl or tricyclo-C₄₋₁₀-alkyl; andwherein, when R¹ is C₃₋₈cycloalkyl, bicyclo-C₄₋₁₀alkyl,tricyclo-C₄₋₁₀-alkyl or aryl, R¹ may optionally be substituted with oneor more substituents selected from halogen, hydroxy, cyano, nitro.C₁₋₆alkyl, C₂₋₆alkenyl, C₂₋₆alkynyl, C₃₋₈cycloalkyl, C₄₋₈cycloalkenyl,C₁₋₆alkoxy, C₁₋₆haloalkoxy and C₁₋₆haloalkyl; R², and R⁴ independentlyrepresent hydrogen, halogen, C₁₋₆alkyl, C₂₋₆alkenyl, C₂₋₆alkynyl,C₃₋₈cycloalkyl, C₃₋₈cycloalkenyl or C₁₋₆alkoxy;

-   -   at least one of R⁵, R⁶ and R⁷ represents C₁₋₆haloalkoxy, and the        remaining of R⁵, R⁶ and R⁷ independently represent hydrogen,        nitro, cyano, halogen, C₁₋₆alkyl, C₂₋₆alkenyl, C₂₋₆alkynyl,        C₃₋₈cycloalkyl, C₄₋₈cycloalkenyl, C₁₋₆haloalkyl, —OR¹⁰,        —NR¹⁰R¹¹, —C(O)OR¹⁰, —COR¹⁰, —C(O)NR¹⁰R¹¹, —SH, —S(O)₂OR¹⁰,        —S(O)₂NR¹⁰R¹¹, —S(O)_(n)R¹¹, aryl or heteroaryl, wherein said        aryl or heteroaryl may optionally be substituted with one or        more C₁₋₆alkyl, halogen, hydroxy or phenyl, R³ represents        hydrogen, halogen, cyano, —OR¹⁰, —NR¹⁰R¹¹, —C(O)OR¹⁰, —COR¹⁰,        —(O)NR¹⁰R¹¹, —S(O)_(n)R¹⁰, —S(O)₂NR¹⁰R¹¹, —NHCOR¹⁰ or —NHSO₂R¹⁰;

n is 0, 1 or 2; and

each R¹⁰ and R¹¹ are selected independently from the group consisting ofhydrogen, C₁₋₆alkyl, C₂₋₆alkenyl, C₂₋₆alkynyl, C₃₋₈cycloalkyl.C₄₋₈cycloalkenyl, C₁₋₆ haloalkyl and C₁₋₆ haloalkoxy; andpharmaceutically acceptable salts, solvates and prodrugs thereof.

Further examples of niclosamide analogues include, but are not limitedto those delineated in Table 4.

TABLE 45-Chloro-N-(4-cyano-2-trifluoromethoxy-phenyl)-3-(1,1-dimethyl-propyl)-2-hydroxy-benzamide3-tert-Butyl-N-(4-cyano-2-trifluoromethoxy-phenyl)-2-hydroxy-5-methanesulfonyl-6-methyl-benzamide3-Bromo-5-tert-butyl-N-(4-cyano-2-trifluoromethoxy-phenyl)-6-hydroxy-2-methyl-benzamide5-Bromo-3-tert-butyl-N-(4-cyano-2-trifluoromethoxy-phenyl)-2-hydroxy-benzamide5-Chloro-3-tert-butyl-N-(4-cyano-2-trifluoromethoxy-phenyl)-2-hydroxy-6-methyl-benzamide3-tert-Butyl-N-(4-cyano-2-trifluoromethoxy-phenyl)-5-fluoro-2-hydroxy-6-methyl-benzamide3-tert-Butyl-N-(4-cyano-2-trifluoromethoxy-phenyl-2-hydroxy-5-methoxy-6-methyl-benzamide3-tert-Butyl-N-(4-cyano-2-trifluoromethoxy-phenyl)-6-ethyl-2-hydroxy-5-methoxy-benzamide 3-tert-Butyl-N-(4-cyano-2-trifluoromethoxy-phenyl)-5-ethanesulfonylamino-2-hydroxy-6-methyl-benzamide3-tert-Butyl-N-(4-cyano-2-trifluoromethoxy-phenyl)-2-hydroxy-6-methyl-5-(propane-1-sulfonylamino)-benzamide3-tert-Butyl-N-(4-cyano-2-trifluoromethoxy-phenyl)-2-hydroxy-6-methyl-5-(propane-2-sulfonylamino)-benzamide3-tert-Butyl-5-cyano-N-(4-cyano-2-trifluoromethoxy-phenyl)-2-hydroxy-6-methyl-benzamide3-Acetyl-5-tert-butyl-N-(4-cyano-2-trifluoromethoxy-phenyl)-6-hydroxy-2-methyl-benzamide3-tert-Butyl-N-(4-cyano-2-trifluoromethoxy-phenyl)-2-hydroxy-5-methanesulfinyl-6-methyl-benzamide3-tert-Butyl-N-(4-cyano-2-trifluoromethoxy-phenyl)-2-hydroxy-6-methyl-5-methylsulfanyl-benzamide3-tert-Butyl-N-(4-cyano-2-trifluoromethoxy-phenyl)-2-hydroxy-5-methanesulfonylamino-6-methyl-benzamide; and3-Acetylamino-5-tert-butyl-N-(4-cyano-2-trifluoromethoxy-phenyl)-6-hydroxy-2-methyl-benzamide

In certain embodiments, the chemical entity can be a compound havingformula XXVII:

where X is N or CR¹⁰; Y is N or CR¹¹; Z is N or CR¹²; and each of R¹,R², R⁵, R⁶, R⁷, R⁸, R⁹, R¹⁰, R¹¹ and R¹² is independently selected fromH, halide (F, Cl, Br, or I), NO₂, OH, OR¹³, SR¹⁴, NR¹⁵R¹⁶, CN, CF₃,acyl, C₁₋₁₀ alkyl, C₂₋₁₀alkenyl, C₂₋₁₀ alkynyl, C₂₋₆ heterocyclyl, C₆₋₁₂aryl, C₇₋₁₄ alkaryl, C₃₋₁₀ alkheterocyclyl, C₁₋₁₀ heteroalkyl, or isdescribed by one of the following formulas:

In compounds of formula XXVII, R³ and R⁴ are independently selected fromthe group consisting of C═O, C═S, C═NR⁴², NH, NR⁴³, CHOR⁴⁴, C₁₋₆alkylene (e.g., CH₂), S═O, S(O)₂, NHC₁₋₆ alkylene (e.g., NHCH₂),C₁₋₆alkyleneNH (e.g., CH₂NH), C₁₋₆ alkylene NR⁴³, NHC(O), C(O)NH. GroupsR² and R⁴; X and R⁴; R⁵ and R³; R⁹ and R³ may combine to form asix-membered ring, using connections described by one of the groups:

For compounds of formula I, each E¹ is independently O, S, or NR⁴²; eachE² is independently CR⁴⁹R⁵⁰, O or S; each E³ is independently CR⁵¹R⁵²,O, S, or NR⁵³; each Q is, independently, O, S, or NR⁵⁴. R¹³ and R¹⁴ areeach independently, acyl, C(O)OC₁₋₆ alkyl, C₁₋₁₀ alkyl, C₂₋₁₀ alkenyl,C₂₋₁₀ alkynyl, C₂₋₆ heterocyclyl. C₆₋₁₂ aryl, C₇₋₁₄ alkaryl, C₃₋₁₀alkheterocyclyl, C₁₋₁₀ heteroalkyl, R¹⁸, R²³, R²⁸, R²⁹, R³⁰, R⁴², R⁵⁴are each, independently, H, C₁₋₁₀ alkyl, C₂₋₁₀ alkenyl, C₂₋₁₀ alkynyl,C₂₋₆ heterocyclyl, C₆₋₁₂ aryl, C₇₋₁₄ alkaryl, C₃₋₁₀ alkheterocyclyl,C₁₋₁₀ heteroalkyl; R¹⁵, R¹⁶, R¹⁷, R¹⁹, R²⁰, R²¹, R²², R²⁴, R²⁵, R²⁶,R²⁷, R⁴³, R⁴⁴, R⁴⁵, R⁴⁶, R⁴⁷, R⁴⁸, R⁵¹, R⁵², and R⁵³ are each,independently, H, C₁₋₁₀ alkyl, C₂₋₁₀alkenyl, C₂₋₁₀ alkynyl, C₂₋₁₀heterocyclyl, C₆₋₁₂ aryl, C₇₋₁₄ alkaryl, C₃₋₁₀ alkheterocyclyl, C₁₋₁₀heteroalkyl; R³¹, R³², R³³, R³⁴, R³⁵, R³⁶, R³⁷, R³⁸, R³⁹, R⁴⁰, R⁴¹, R⁴⁹,and R⁵⁰ are each, independently, H, halide, NO₂, CN, CF₃, C₁₋₁₀ alkyl,C₂₋₁₀ alkenyl, C₂₋₁₀ alkynyl, C₂₋₆ heterocyclyl, C₆₋₁₂ aryl, C₇₋₁₄alkaryl, C₃₋₁₀ alkheterocyclyl, or C₁₋₁₀ heteroalkyl.

Further examples of niclosamide analogues include, but are not limitedto those delineated in Table 5.

TABLE 5

In certain of the foregoing embodiments, acyl is a chemical moiety withthe formula R—C(O)—, where R is selected from C₁₋₁₀ alkyl, C₂₋₆heterocyclyl (e.g., heteroaromatic), and C₆₋₁₂ aryl.

Further examples of niclosamide analogues include, but are not limitedto those delineated in Table 6.

TABLE 6

In certain embodiments, the chemical entity is niclosamide or apharmaceutically acceptable salt or hydrate thereof. “Niclosamide”refers to a compound having the following chemical structure:

Niclosamide is known by the IUPAC designation:2′5-dichloro-4′-nitrosalicylanilide and by the CAS designation. CAS:5-chloro-N-(2-chloro-4-nitrophenyl)-2-hydroxybenzamide. Niclosamide hasa relatively low water solubility at about from 5-8 mg/L at 20° C., issparingly soluble in ether, ethanol and chloroform, and is soluble inacetone. The ethanolamine salt dissolves in distilled water 180-280mg/L, at 20° C.

Niclosamide is available in a various salt or solvated forms. Theseinclude, but are not limited to, the ethanolamine salt known by theIUPAC designation 5-chloro-salicyl-(2-chloro-4-nitro) anilide2-aminoethanol salt or the CAS designation5-chloro-N-(2-chloro-4-nitrophenyl)-2-hydroxybenzamide with2-aminoethanol (1.1)—see, e.g., US 2013/0231312; the piperazine saltknown by the IUPAC designation 5-chloro-salicyl-(2-chloro-4-nitro)anilide piperazine salt or the CAS designation5-chloro-N-2-chloro-4-nitrophenyl)-2-hydroxybenzamide with piperazine(2:1); and niclosamide monohydrate known by the IUPAC designation5-chloro-salicyl-(2-chloro-4-nitro) anilide monohydrate or the CASdesignation 5-chloro-N-(2-chloro-4-nitrophenyl)-2-hydroxybenzamide withmonohydrate (1:1).

Niclosamide is commercially available in a variety of formulationsincluding, but not limited to BAYER 73®, BAYER 2353®, BAYER 25 648®,BAYLUSCID®, BAYLUSCIDE®, CESTOCID®, CLONITRALID, DICHLOSALE®, FENASAL®,HL 2447®, IOMESAN®, IOMEZAN®, LINTEX®, MANOSIL®, NASEMO®, NICLOSAMID®,PHENASAL®, TREDEMINE®, SULQUI®, VERMITID®, VERMITIN®, YOMESAN®, and thelike.

Compounds disclosed herein are commercially available or can be readilyprepared from commercially available starting materials according toestablished methodology in the art of organic synthesis. General methodsof synthesizing the compound can be found in, e.g., Stuart Warren andPaul Wyatt, Workbook for Organic Synthesis: The Disconnection Approach,second Edition, Wiley, 2010. See also, e.g., U.S. Pat. No. 8,148,328,which is incorporated herein by reference in its entirety and Mook, etal., Bioorg. Med. Chem 2015, 23, 5829, which is incorporated herein byreference in its entirety.

In other embodiments, the chemical entity can be selected from thecompounds that are disclosed generically, sub generically andspecifically in any one or more of WO 2004/006906; WO 2006/120178; US2009/0062396; WO 2012/143377: WO 2012/068274; U.S. Pat. Nos. 7,132,546;7,989,498; and 8,263,857; each of which is incorporated herein byreference in its entirety.

Other Chemical Entities

In some embodiments, the chemical entity can be an anthelminthic agentselected from nitazoxanide, closantel, pyrvinium pamoate, andsalinomycin. See. e.g., Senkowski, W., et al., Mol. Cancer Ther. 2015,14, 1504.

Cocrystals of Chemical Entities

Overview

In some embodiments, the chemical entity can be in the form of acocrystal that includes (i) the chemical entity (e.g., a mitochondrialuncoupling agent (e.g., niclosamide or a niclosamide analogue) or apharmaceutically acceptable salt and/or hydrate thereof; and (ii) one ormore pharmaceutically acceptable coformers. The term “co-crystal” asused herein refers to a crystalline material comprised of two or moreunique solids at room temperature in a stoichiometric ornon-stoichiometric ratio, which are held together in the crystal latticeby one or more non-covalent interactions (e.g., hydrogen bonds,pi-stacking, guest-host complexation and van der Waals interactions).

In some embodiments, at least one of the one or more non-covalentinteractions is a hydrogen bond. In certain of these embodiments, thechemical entity is the hydrogen bond donor, and one of one or morecoformers is the hydrogen bond acceptor. In other embodiments, thechemical entity is the hydrogen bond acceptor, and one of one or morecoformers is the hydrogen bond donor.

The co-crystals described herein can include one or more solvate (e.g.,water or an organic solvent containing one or more hydroxyl groups,e.g., a C₁-C₆ alcohol or diol, e.g., a C₁-C₆ alcohol or diol, e.g.,ethanol or propylene glycol) molecules in the crystalline lattice.However, solvates of chemical entities that do not further comprise acoformer (e.g., a solid conformer) are not encompassed by the co-crystaldefinition set forth in this disclosure.

In some embodiments, the cocrystal includes more than one coformer. Forexample, two, three, four, five, or more co formers can be incorporatedin a co-crystal with the chemical entity. The ratio of the chemicalentity to each of the one or more pharmaceutically acceptable coformersmay be stoichiometric or non-stoichiometric. As a non-limiting example,1:1, 1:1.5 and 1:2 ratios of chemical entity:coformer are contemplated.

The chemical entity and each of the one or more pharmaceuticallyacceptable coformers may each be independently specified as a free form,or more specifically, a free acid, free base, or zwitter ion; a salt, ormore specifically for example, an inorganic base addition salt such assodium, potassium, lithium, calcium, magnesium, ammonium, aluminum saltsor organic base addition salts, or an inorganic acid addition salts suchas HBr, HCl, sulfuric, nitric, or phosphoric acid addition salts or anorganic acid addition salt such as acetic, proprionic, pyruvic, malanic,succinic, malic, maleic, fumaric, tartaric, citric, benzoic,methanesulfonic, ethanesulforic, stearic or lactic acid addition salt;an anhydrate or hydrate of a free form or salt, or more specifically,for example, a hemihydrate, monohydrate, dihydrate, trihydrate,quadrahydrate, pentahydrate; or a solvate of a free form or salt.

The Chemical Entity

In some embodiments, the chemical entity (e.g., a mitochondrialuncoupling agent (i.e., component (i) above) can form one or morehydrogen bonds with the one or more pharmaceutically acceptablecoformers in the cocrystal. In some embodiments, the chemical entity canaccept one or more hydrogen bonds from the one or more pharmaceuticallyacceptable coformers in the cocrystal. In some embodiments, the chemicalentity can form one or more hydrogen bonds with the one or morepharmaceutically acceptable coformers, and the chemical entity canaccept one or more hydrogen bonds with the one or more pharmaceuticallyacceptable coformers in the cocrystal.

In some embodiments, the chemical entity (e.g., a mitochondrialuncoupling agent (i.e., component (i) above) includes one or morefunctional groups selected from the group consisting of: ether,thioether, hydroxy, sulfhydryl, aldehyde, ketone, thioketone, nitrateester, phosphate ester, thiophosphate ester, ester, thioester, sulfateester, carboxylic acid, phosphonic acid, phosphinic acid, sulfonic acid,amido, primary amine, secondary amine, ammonia, tertiary amino, sp2amino, thiocyanate, cyanamide, oxime, nitrile, diazo, haloalkyl, nitro,heterocyclic ring, heteroaryl ring, epoxide, peroxide, and hydroxamicacid.

In some embodiments, the chemical entity (e.g., a mitochondrialuncoupling agent (i.e., component (i) above) is niclosamide or apharmaceutically acceptable salt or hydrate thereof; or a niclosamideanalog, or a pharmaceutically acceptable salt or hydrate thereof. Insome of these embodiments, the chemical entity can be a compound havingany one of formulas (I) and (XVIII)-(XXV), e.g., formula XXIV, XXV, orXXVII; or any one of the list of coformers delineated below. In certainof these embodiments, the chemical entity can be a niclosamide analoguehaving any one of formulas (I) and (XVIII)-(XXV), e.g., formula XXIV orXXV; or XXVI; or any one of the list of coformers delineated below. Incertain of these embodiments, the chemical entity can be a niclosamideor a pharmaceutically acceptable salt or hydrate thereof (e.g.,niclosamide).

Coformers

In some embodiments, at least one of the one or more pharmaceuticallyacceptable coformers can form one or more hydrogen bonds with thechemical entity in the cocrystal. In some embodiments, at least one ofthe one or more pharmaceutically acceptable coformers can accept one ormore hydrogen bonds from the chemical entity in the cocrystal. In someembodiments, at least one of the one or more pharmaceutically acceptablecoformers can form one or more hydrogen bonds with the chemical entityin the cocrystal, and at least one of the one or more pharmaceuticallyacceptable coformers can accept one or more hydrogen bonds from thechemical entity in the cocrystal.

In some embodiments, at least one of the one or more pharmaceuticallyacceptable coformers comprises one or more functional groups selectedfrom the group consisting of: ether, thioether, hydroxy, sulfhydryl,aldehyde, ketone, thioketone, nitrate ester, phosphate ester,thiophosphate ester, ester, thioester, sulfate ester, carboxylic acid,phosphonic acid, phosphinic acid, sulfonic acid, amido, primary amine,secondary amine, ammonia, tertiary amino, sp2 amino, thiocyanate,cyanamide, oxime, nitrile, diazo, haloalkyl, nitro, heterocyclic ring,heteroaryl ring, epoxide, peroxide, and hydroxamic acid.

In certain embodiments, each of the one of the one or morepharmaceutically acceptable coformers is independently selected fromacetamide, benzamide, (+/−)-limonene, 1-(phenylazo)-2-naphthylamine,1,2,6-hexanetriol,1,2-dimyristoyl-sn-glycero-3-(phospho-s-(1-glycerol)),1,2-dimyristoyl-sn-glycero-3-phosphocholine,1,2-dioleoyl-sn-glycero-3-phosphocholine,1,2-dipalmitoyl-sn-glycero-3-(phospho-rac-(1-glycerol)),1,2-distearoyl-sn-glycero-3-(phospho-rac-(1-glycerol)),1,2-distearoyl-sn-glycero-3-phosphocholine, 1,5-naphthalene-disulfonicacid, 1-hydroxy-2-naphthoic acid, 1-o-tolylbiguanide,2-ethyl-1,6-hexanediol, 4-aminobenzoic acid, 4-aminopyridine,4-aminosalicylic acid, 4-chlorobenzene-sulfonic acid, 4-ethoxyphenylurea, 7-oxo-dhea, acacia, acacia mucilage, acacia syrup, acesulfame,acesulfame potassium, acetohydroxamic acid, acetone sodium bisulfite,acetylated lanolin alcohols, acetylated monoglycerides, acetylcysteine,acetyltributyl citrate, acrylates copolymer, acrylic acid-isooctylacrylate copolymer, adenine, adipic acid, alanine, albumin aggregated,albumin colloidal, albumin human, albumins, alginic acid, alkyl ammoniumsulfonic acid betaine, alkyl aryl sodium sulfonate, allantoin,allopurineol, allyl alpha-ionone, alpha-terpineol, alpha-tocopherol,alpha-tocopherol acetate, aminobenzoate sodium, amyl acetate, anethole,anhydrous citric acid, anhydrous dextrose, anhydrous lactose, anhydroustribasic sodium phosphate, anhydrous trisodium citrate, arginine,arlacel, asafetida, ascorbic acid, ascorbyl palmitate, asparagine,aspartame, aspartic acid, bacteriostatic sodium chloride injection,barium sulfate, benzalkonium chloride, benzenesulfonic acid,benzethonium chloride, benzododecinium bromide, benzoic acid, benzylacetate, benzyl alcohol, benzyl benzoate, benzyl chloride,beta-carotene, betanaphthol, betose, bibapcitide, bismuth subcarbonate,bismuth subgallate, boric acid, brocrinat, butyl stearate, butylatedhydroxyanisole, butylated hydroxytoluene, butylparaben, butyric acid,C-11-1-aminocyclohexanecarboxylic acid, C12-15 alkyl lactate, caffeine,calcobutrol, caldiamide sodium, caloxetate trisodium, calteridolcalcium, camphoric acid, capric acid, captan, captisol,carboxypolymethylene, carmine, carnauba wax, carnauba yellow wax,carrageenan, carrageenan calcium, carrageenan salt, carrageenan sodium,ceresin, ceteareth-12, ceteareth-15, ceteareth-30, cetearylalcohol/ceteareth-20, cetearyl ethylhexanoate, ceteth-10, ceteth-2,ceteth-20, ceteth-23, cetostearyl alcohol, cetrimonium chloride, cetylalcohol, cetyl esters wax, cetyl palmitate, cetylpyridinium chloride,chlorocresol, chloroxylenol, cholesterol, chrysin, cinnamaldehyde,cinnamic acid, citrate, citric acid, citric acid monohydrate, clemizole,cocamide ether sulfate, cocamine oxide, coco betaine, cocodiethanolamide, coco monoethanolamide, coco-caprylate, coco-glycerides,creatine, creatinine, cresol, cupric sulfate, cyclamic acid,cyclomethicone, cyclomethicone 5, cysteine, dalfampridine, decyl methylsulfoxide, dehydroacetic acid, denatonium benzoate, deoxycholic acid,dextran, dextran 40, dextrates, dextrin, dextrose, dextrose monohydrate,diacetylated monoglycerides, diatrizoic acid, dibasic anhydrous sodiumphosphate, dibasic sodium phosphate, dibasic sodium phosphate dihydrate,dibasic sodium phosphate dodecahydrate, dibasic sodium phosphateheptahydrate, dibutyl phthalate, dibutyl sebacate, diethyl phthalate,diethyl pyrocarbonate, diethyl sebacate, diethylaminoethyl stearamidephosphate, diethylene glycol monoethyl ether, diethylene glycolmonomethyl ether, diethylhexyl phthalate, diisopropyl adipate,diisopropyl dilinoleate, diisopropylbenzothiazyl-2-sulfenamide,dimethicone medical fluid 360, dimethyl isosorbide, dimethyl phthalate,dimethyl sulfoxide, dimethyldioctadecylammonium bentonite,dimethylglycine, dimethylsiloxane/methylvinylsiloxane copolymer,dinoseb-ammonium, dipropylene glycol, disodium cocoamphodiacetate,disodium hydrogen citrate, disodium laureth sulfosuccinate, disodiumlauryl sulfosuccinate, disodium oleamido monoethanolaminesulfosuccinate, disodium sulfosalicylate, disofenin, dl-a350 lacticacid, dl-acetyltryptophan, dl-alpha-tocopherol, dl-alpha-tocopherolacetate, dl-dipalmitoylphosphatidylglycerol,dl-distearoylphosphatidylcholine, dl-glutamic acid, dl-tartaric acid,d-mannose, dmdm hydantoin, docosanol, docusate sodium, d-ribose, edetatecalcium disodium, edetate disodium, edetate sodium, edetic acid, eggphosphatidyl glycerol, egg phospholipids, entsufon, entsufon sodium,epilactose, epitetracycline hydrochloride, erythorbic acid, erythritol,ethanolamine hydrochloride, ethyl maltol, ethyl oleate, ethyl vanillate,ethyl vanillin, ethylenediamine dihydrochloride, ethylhexylhydroxystearate, ethylparaben, eucalyptol, eugenol, exametazime, fattyacid esters, fatty acid glycerides, fatty acid pentaerythriol ester,fatty acids, fatty alcohol citrate, fatty alcohols, ferric chloride,ferric oxide, ferrosoferric oxide, ferrous fumarate, ferrous oxide,fluorescein, fructose, fumaric acid, fumaryl diketopiperazine,gadolinium oxide, galactaric acid, galactose, gamma cyclodextrin,genistein, gentisic acid, gentisic acid ethanolamide, gentisic acidethanolamine, gluceptate sodium, gluconic acid, gluconolactone,glucosamine, glucose, glucuronic acid, glutamic acid, glutamic acidhydrochloride, glutamine, glutaric acid, glutathione, glycerylcaprylate, glyceryl dibehenate, glyceryl distearate, glycerylisostearate, glyceryl laurate, glyceryl monostearate, glyceryl oleate,glyceryl palmitate, glyceryl palmitostearate, glyceryl ricinoleate,glyceryl stearate, glyceryl stearate-laureth-23, glyceryl stearate/pegstearate, glyceryl stearate/peg-100 stearate, glyceryl stearate/peg-40stearate, glyceryl stearate-stearamidoethyl diethylamine, glyceryltrioleate, glycine, glycine hydrochloride, glycol distearate, glycolstearate, glycolic acid, glycyrrhizin, guanidine hydrochloride,hexylresorcinol, hippuric acid, histidine, hyaluronate sodium,hydrocortisone, hydroquinone, hydrous-citric acid,hydroxyethylpiperazine ethane sulfonic acid, hydroxyoctacosanylhydroxystearate, hydroxyprogesterone caproate, hydroxypropylbeta-cyclodextrin, hystrene, illicium anisatum, imidazole, imidurea,indigotindisulfonate sodium, iodoxamic acid, iofetamine hydrochloride,ipriflavone, isoleucine, isopropyl isostearate, isopropyl myristate,isopropyl myristate-myristyl alcohol, isopropyl palmitate, isopropylstearate, isostearic acid, isostearyl alcohol, lactate, lactitolmonohydrate, lactobionic acid, lactose, landalgine, lanolin,lauralkonium chloride, lauramine oxide, laureth sulfate, lauric acid,lauric diethanolamide, lauric myristic diethanolamide, lauroylsarcosine, lauryl lactate, lauryl sulfate, lecithin, leucine,levomenthol, levulinic acid, lidofenin, 1-sodium lactate, lysine, maleicacid, malic acid, malonic acid, maltitol, maltodextrin, maltol, maltoseanhydrous, mandelic acid, mannitol, maprofix, mebrofenin, medium-chaintriglycerides, medronate disodium, medronic acid, menthol, metacresol,methionine, methyl salicylate, methyl stearate,methylchloroisothiazolinone, methylisothiazolinone, methylparaben,methylparaben sodium, miripirium chloride, mono and diglyceride,monobasic sodium phosphate, monobasic sodium phosphate anhydrous,monobasic sodium phosphate dihydrate, monobasic sodium phosphatemonohydrate, monoglyceride citrate, monoglycerides, monosodium citrate,monosodium glutamate, monostearyl citrate, monothioglycerol, myristicacid, myristyl alcohol, myristyl lactate, niacinamide, nicotinamide,nicotinic acid, N-methyl glucamine, octanoic acid, oleth-20, oleylalcohol, oleyl oleate, orotic acid, oxalic acid, oxidronate disodium,oxyquinoline, palmitamine oxide, palmitic acid, pamoic acid,pentadecalactone, pentaerythritol cocoate, pentasodium pentetate,pentetate calcium trisodium, pentetic acid, phenol, phenonip,phenoxyethanol, phenylalanine, phenylethyl alcohol, phospholipid,piperazine, piperazine hexahydrate, procaine, product wat, proline,propenyl guaethol, propyl gallate, propylene carbonate, propyleneglycol, propylene glycol-lecithin, propylene glycol alginate, propyleneglycol diacetate, propylene glycol dicaprylate, propylene glycolmonolaurate, propylene glycol monopalmitostearate, propylene glycolpalmitostearate, propylene glycol ricinoleate, propyleneglycol/diazolidinyl urea/methylparaben/propylparben, propylparaben,propylparaben sodium, p-toluenesulfonic acid, pyridoxamine, pyridoxine(4-pyridoxic acid), quercetin, resveratrol, riboflavin, saccharin,saccharin calcium, saccharin sodium, saccharin sodium anhydrous,salicylic acid, saturated fatty acid esters, sebacic acid, serine,sodium 1,2-ethanedisulfonate, sodium 2-naphthalenesulfonate, sodiumacetate, sodium acetate anhydrous, sodium alginate, sodium alkylsulfate, sodium aluminium silicate, sodium ascorbate, sodium benzoate,sodium bicarbonate, sodium bisulfate, sodium bisulfate acetone, sodiumbisulfite, sodium bitartrate, sodium borate, sodium borate decahydrate,sodium carbonate, sodium carbonate decahydrate, sodium carbonatemonohydrate, sodium carboxymethyl beta-glucan (ds 065-085), sodiumcaseinate, sodium cellulose, sodium cetostearyl sulfate, sodiumchlorate, sodium chloride, sodium chloride injection, sodium cholesterylsulfate, sodium citrate, sodium citrate hydrous, sodium cocoylsarcosinate, sodium cyclamate, sodium desoxycholate, sodium dithionite,sodium dodecylbenzenesulfonate, sodium ethylparaben, sodium formaldehydesulfoxylate, sodium gluconate, sodium hydroxide, sodium hypochlorite,sodium iodide, sodium lactate, sodium laureth-2 sulfate, sodiumlaureth-3 sulfate, sodium laureth-5 sulfate, sodium lauroyl sarcosinate,sodium lauryl sulfate, sodium lauryl sulfoacetate, sodium metabisulfite,sodium nitrate, sodium oleate, sodium phosphate, sodium phosphatedihydrate, sodium phosphite, sodium polyacrylate, sodium polyacrylate(2500000 MW), sodium polymetaphosphate, sodium propionate, sodiumpyrophosphate, sodium pyrrolidone carboxylate, sodium starch glycolate,sodium starch glycolate type a corn, sodium starch glycolate type apotato, type B potato sodium starch glycolate, sodium stearate, sodiumstearyl fumarate, sodium succinate hexahydrate, sodium sulfate, sodiumsulfate anhydrous, sodium sulfate decahydrate, sodium sulfite, sodiumsulfosuccinated undecyclenic monoalkylolamide, sodium tartrate, sodiumthioglycolate, sodium thiomalate, sodium thiosulfate, sodium thiosulfateanhydrous, sodium trimetaphosphate, sodium tripolyphosphate, sodiumxylenesulfonate, sorbic acid, sorbitan, sorbitan isostearate, sorbitanmonolaurate, sorbitan monooleate, sorbitan monopalmitate, sorbitanmonostearate, sorbitan sesquioleate, sorbitan trioleate, sorbitantristearate, sorbitol, squalane, stannous 2-ethylhexanoate,stearalkonium chloride, stearalkonium hectorite/propylene carbonate,stearamidoethyl diethylamine, stearates, stearic acid, stearicdiethanolamide, stearoxytrimethylsilane, stearyl alcohol, succinic acid,sucralose, sucrose, sucrose distearate, sucrose laurate, sucrosepalmitate, sucrose polyesters, sucrose stearate, sucrose syrup,sulfacetamide sodium, sulfobutylether beta-cyclodextrin, tagatose,tartaric acid, tegacid, tert-butylhydroquinone, tetrofosmin,theophylline, thimerosal, threonine, thymol, tocopherol, tocophersolan,tragacanth, triacetin, tribasic sodium phosphate, tribasic sodiumphosphate monohydrate, tribehenin, tricaprylin, triceteareth-4phosphate, triethanolamine lauryl sulfate, triethyl citrate,trihydroxystearin, trilaneth-4 phosphate, trilaureth-4 phosphate,trimyristin, tris, trisodium citrate dihydrate, trisodium hedta,tristearin, trolamine, tromantadine, tromethamine, tryptophan,tyloxapol, tyrosine, undecylenic acid, urea, urethane, ursodiol, valine,vanillin, versetamide, viscarin, vitamin E, vitamin E acetate, vitaminK5, xylitol, and zinc sulfate. See also U.S. Pat. No. 7,927,613, whichis incorporated herein by reference in its entirety. Otherpharmaceutically acceptable coformers include those delineated in the“Generally Regarded as Safe” (“GRAS”) and/or the US FDA “EverythingAdded to Food in the United States” (“EAFUS”) lists.

In certain embodiments, at least one of the one or more pharmaceuticallyacceptable coformers is selected from the group consisting of caffeine,urea, p-aminobenzoic acid, theophylline, benzyl benzoate, andnicotinamide. In other embodiments, the one or more pharmaceuticallyacceptable coformers is other than those selected from the groupconsisting of caffeine, urea, p-aminobenzoic acid, theophylline, benzylbenzoate, and nicotinamide. In other embodiments, the one or morepharmaceutically acceptable coformers is other than those selected fromthe group consisting of acetamide, benzamide, 2-aminothiazole, andisoniazide. In still other embodiments, the one or more pharmaceuticallyacceptable coformers is an amino acid (e.g., proline, e.g., D-proline orL-proline, or racemic proline). In another embodiment, the one or morepharmaceutically acceptable coformers is a 5-10 (e.g., 5-9, 5-6, or 5)membered heteroaryl, e.g., a nitrogen-containing heteroaryl, e.g.,imidazole.

In certain embodiments, at least one of the one or more pharmaceuticallyacceptable coformers is a second API. In certain of these embodiments,the second API is independently selected from (−)-amlodipine,(−)-halofenate, (R)-salbutamol, (R)-salbutamol, (R,R)-formoterol,(S)-doxazosin, (S)-fluoxetine, (S)-oxybutynin, 1,2-naphthoquinone,17-methyltestosterone, 17α-hydroxyprogesterone, 195mPt-cisplatin,1-naphthyl salicylate, 1-naphthylamine-4-, 1-theobromineacetic,1α-hydroxycholecalciferol, 2,4,6-tribromo-m-cresol,2,6-diamino-2′-butyloxy-3,5′-azopyridine,2-[[[(1r)-2-(1h-imidazol-4-yl)-1-methylethyl]imino]phenylmethyl]-phenol,21-acetoxypregnenolone, 2-amino-4-picoline, 2-aminothiazole,2-ethoxybenzoic acid, 2-naphthol, 2-naphthyl benzoate, 2-naphthyllactate, 2-naphthyl salicylate, 2-p-sulfanilylanilinoethanol,2-thiouracil, 3′,3″,5′,5″-tetra-bromophenolphthalein,3-amino-4-hydroxybutyric acid, 3-Bromo-D-camphor, 3-Hydroxycamphor,3-O-Lauroylpyridoxol Diacetate, 3-pentadecylcatechol, 3-quinuclidinol,4,4′-oxydi-2-butanol, 4,4′-sulfinyldianiline, 4-amino-3-hydroxybutyricacid, 4-amino-3-phenylbutyric acid, 4-aminosalicylic acid,4-chloro-m-cresol, 4-hexylresorcinol, 4-salicyloylmorpholine,5′-nitro-2′-propoxyacetanilide, 5-aminolevulinic acid, 5-azacitidine,5-bromosalicyl-hydroxamic acid, 5F-DF-203, 5-FU, 5-HT3 antagonists,6-azauridine, 6-mercaptopurine, 8-hydroxyquinoline, 9-aminocamptothecin,A-151892, A-5021, abacavir, abaperidone, abarelix, abciximab, abecarnil,abetimus, abiraterone, ABLC, ABT-751, AC-5216, acadesine, acamprosate,acamprosate, acarbose, acebrophylline, acebutolol, acecainide,acecarbromal, aceclofenac, acedapsone, acediasulfone, acefylline,aceglutamide, aceglutamide, acemetacin, acenocoumarol, aceponate,acetal, acetamidoeugenol, acetaminophen, acetaminosalol, acetanilide,acetarsone, acetazolamide, acetiamine, acetohexamide, acetohydroxamicacid, acetophenazine, acetophenide, acetophenone, acetosulfone,acetoxolone, acetrizoat, acetyl, acetylcarnitine, acetylcholine,acetylcholine, acetylcysteine, acetylleucine, acetylpheneturide,acetylsalicylate, acetylsalicylic acid, aciclovir, acifran, acipimox,acitazanolast, acitretin, aclarubicin, aclatonium, aconitine, Acranil®,acriflavine, acrisorcin, acrivastine, acrivastine, actagardinederivative, actarit, ACTH, acyclovir, adapalene, ADCON-L, adefovir,adefovir dipivoxil, adenoscan, adenosine triphosphate, ADEPT,adinazolam, adiphenine, ADL-10-0101, adrafinil, adrenalone,adrenochrome, adrogolide, AEOL-10150, aesthinol, AET, AF-2259,afloqualone, AG-041R, AG-2037, AGN-194310, agomelatine, ahistan,AHL-157, AIT-034, AIT-202, AJ-9677, AJG-049, ajmaline, akzo desogestrel,alacepril, alapivoxil, albaconazole, albendazole, albuterol, albutoin,alclofenac, alclometasone, alcuronium, aldioxa, aldol, aldosterone,alendronate, alendronic acid, alexidine, alfacalcidol, alfadolone,alfaxalone, alfentanil, alfimeprase, alfuzosin, alfuzosin, algestone,algestone, algin, alglucerase, alibendol, aliskiren, alitertinoin,alizapride, alkannin, alkofanone, allantoin, allobarbital, allopurinol,allyl isothiocyanate, allylestrenol, almagate, alminoprofen, almitrine,almotriptan, aloe-emodin, aloin, alosetron, alovudine, aloxiprin,alpha-, alpha-1 protease, alphaprodine, alpidem, alpiropride,alprazolam, alprenolol, alsactide, ALT-711, Althiazid, altinicline,altretamine, aluminium chloride hexahydrate, aluminon, aluminum acetatesolution, aluminum chlorate, aluminum hydroxychloride, aluminumpotassium sulfate, aluminum sodium sulfate, alusulf, alverine,alvimopan, alvocidib, ALX-0646, AM-24, AM-36, AM-477, amantadine,amantanium, ambazon, ambenonium, ambrisentan, ambroxol, ambucaine,ambuphylline, ambusid, ambutonium bromide, amcinonide, AMD-3100,amdinocillin, amdinocillin pivoxil, amdoxovir, amelubant, americaine,amezinium, amfenac, amidephrine, amidinomycin, amifostine, amiglumide,amikacin, amiloride, aminacrine, amineptine, aminitrozole, amino acidpreparations, aminocaproic acid, aminoglutethimide, aminoguanidine,aminohippurate, aminometradine, aminopentamide, aminophylline,aminopromazine, aminopyrine, aminoquinuride, aminorex, amiodarone,amiodipine, amiphenazole, amiprilose, amisulpride, amitriptyline,amitriptyline+ketamine, amitriptylinoxide, amlexanox, ammoniacum,ammoniated mercuric chloride, ammonium benzoate, ammonium mandelate,ammonium salicylate, ammonium valerate, amobarbital, amocarzine,amodiaquin, amorolfine, amoscanat, amosulalol, amotriphene, amoxapine,amoxicillin, amoxicillin+potassium clavulan, AMPAlex, amphetamine,amphetaminil, amphotericin B, ampicillin, ampiroxicam, ampligen,amprenavir, amrinose, amrubicin, amsacrine, amtolmetin guacil,amylocaine, AN-152, anabolic steroids, anagestone, anagrelide,anastrozole, anazolene, ancitabine, ancrod, andolast, androisoxazole,androstenediol, anecortave, anethole, anethole trithione, angiogenix,angiotensin, anhydrovinblastine, anidulafungin, anilerdine, aniracetam,anisindione, anisomycin, anisotropine, anistreplase, antazoline,anthiolimine, anthralin, anthramycin, anthrarobin, anthrax inhibitor,antiangiogenic, anticort, antidepressants, anti-invasins, antimonypotassium tartrate, antimony sodium thioglycollate, antimonythioglycollamide, antiprogestin, antipyrine, antipyrine salicylate,antithrombin III, anxiolytics, AP-521, AP-5280, apalcillin, apaziquone,apazone, apocodeine, apomine, apomorphine, apraclonidine, aprepitant,aprindine, aprobarbital, apronalide, aprotinin, aptiganel, AQ4N,aquavan, AR-116081, AR-A2, arachidonic acid, aranidipine, arbekacin,arbidol, arbutamine, arcitumomab, ardeparin, arecoline, argatroban,arginine, Ariflo®, aripiprazole, arofylline, arotinolol, arsacetin,arsenic trioxide, arsphenamine, arteether, arteflene, artemether,artemisinin, artemotil, artesunate, arzoxifene, AS-3201, ASA,ascaridole, ascorbic acid, asenapine, asimadoline, asocarboxazid,asoprisnil, asoxime, aspartic acid, aspidin, aspidinol, aspirin, aspirindipyridamole, aspoxicillin, AST-120, astemizole, asulacrine, AT-1015,atamestane, atazanavir, atenolol, atenolol+chlorthalidone,atenolol+nifedipine, atevirdine, atipamezole, atiprimod dimaleate,ATL-146e, atomoxetine, atorvastatin, atosiban, atovaquone,atovaquone+proguanil, atracurium, atrasentan, atrial natriuretic,atrolactamide, atropine, augmentin, auranofin, aurothioglucose,avasimibe, avobenzone, AWD-12-281, azacitidine, azacyclonol,azanidazole, azapropazone, azaserine, azasertron, azatadine,azathipprine, AZD-4282, AZD-6140, azelaic acid, azelastine,azelnidipine, azidamfenicol, azidocillin, azimilide, azintamide,azithromycin, azlocillin, azosemide, aztreonam, azulene, bacampicillin,bacitracin, baclofen, baicalein, balofloxacin, balsalazide, bambuterol,bamethan, bamifylline, bamipine, barbital, barnidipine, BAS-118, basicalumina, baslilximab, batimastat, batroxobin, Bay-41-2272, Bay-41-8543,BAY-43-9006, BAY-57-1293, bazedoxifen, BBR-3464, BBR-3576, BBR-3610,BCH-1868, bebeerine, beclamide, beclometasone, befloxatone, befunolol,bemegride, benactyzine, benazepril, bencyclane, bendazac,bendroflumethiazide, benetonide, benexate, benfluorex, benfotiamine,benfurodil, benidipine, benorylate, benoxaprofen, benoxinate,benperidol, benproperine, benserazide, bentazepam, bentiromide,bentoquatam, benzafibrate, benzalkonium, benzarone, benzathine,benzbromarone, benzethonium, benzetimide, benzilonium, benziodarone,benznidazole, benzocaine, benzoctamine, benzonatate, benzoxoniumchloride, benzoyl peroxide, benzoylpas, benzphetamine, benzpiperylon,benzquinamide, benzthiazide, benztropine, benzydamine, benzyl benzoate,benzylhydrochloro-thiazide, benzylmorphine, bephenium, bepotastine,bepridil, beraprost, berberine, bergapten, bermoprofen, besipirdine,betahistine, betaine, betamethasone, betamipron, betasine, betaxolol,betazole, bethanechol, bethanidine, betoxycaine, bevantolol, bevonium,bexarotene, bezitramide, BG-9928, BIA-2-024, BIA-2-093, BIA-3-202,bialamicol, biapenem, bibenzonium, bibrocathol, bicalutamide,bicifadine, bicisate, bicyclic, bidisomide, bietamiverine, bietanautine,bietaserpine, bifermelane, bifluranol, bifonazole, bimatoprost,bimoclomol, bimosiamose, binifibrate, binodenoson, biomed-101, biotin,biperiden, biriperone, birlcodar, bisacodyl, bisantrene, bisbentiamine,bisdequalinium, bismuth, bismuth, bismuth, bismuth aluminate, bismuthethyl, bismuth sodium, bismuth sodium triglycollamate, bismuthsubcarbonate, bismuth subgallate, bismuth subnitrate, bismuthsubsalicylate, bisoprolol, bisoprolol+HCTZ,bisoprolol+trichloromethiazide, bisoxatin, bithionol, bitolterol,bitoscanat, BL-3875, bleomycin, blonanserin, BMS-184476, BMS-387032,BN-82451, BNP-7787, BO-653, bolandiol, bolasterone, boldenone,bopindolol, bornyl chloride, bornyl salicylate, bortezomib, bosentan,bradycor, brain natriuretic, brallobarbital, brasofensine, brequinar,bretylium, brilliant green, brimonidine, brinzolamide, brivudin,brodimoprim, bromazepam, bromfenac, bromhexine bromide, bromindione,bromisovalum, bromocriptine, bromo-diphenhydramine, bromoform,bromopride, bromo-salicychloranilide, bromperidol, brompheniramine,broparoestrol, bropirimine, brostallicin, brotizolam, brovincamine,broxyquinoline, brozuridine, brucine, bucetin, bucillamine, bucindolol,bucladesine, buclizine, buclosamide, bucolome, bucricaine, bucumolol,budesonide, budesonide+formoterol, budipine, budralazine, bufeniode,bufetolol, bufexamac, buflomedil, buformin, bufuralol, bumadizon,bumetanide, bunaftine, bunamiodyl sodium, bunazosin, bunitrolol,bupivacaine, bupranolol, buprenorphine, bupropion, buramate, buserelin,buspirone, busulfan, busulfan, butabarbital, butacaine, butacetin,butalamine, butalbital, butallylonal, butamben, butamirate,butanilicaine, butaperazine, butaverine, butazolamide, butedronic acid,butenafine, butethal, butethamate, butethamine, buthalital, buthiazide,butibufen, butidrine, butobendine, butoconazole, butoctamide,butofilolol, butorphanol, butoxycaine, butriptyline, butropium,butylthiolaurate, butyrate propio, buzepide, BVT-5182, BXT-51072,C-1311, cabergoline, cabergoline, cacodylic acid, cactinomycin,cadexomer iodine, cadmium salicylate, cadralazine, cafaminol, caffeine,calcifediol, calcipotriene, calcipotriol, calcipotriol+beclometasone,calcitriol, calcium 3-aurothio-2-propanol-1-sulfonate, calciumacetylsalicylate, calcium bromolactobionate, calcium carbonate, calciumgluconate, calcium glycerophosphate, calcium hopantothenate, calciumiodobehenate, calcium iodosterate, calcium lactate, calcium levulinate,calcium mesoxalate, calcium N-carbamoylaspartate, calcium polycarbophil,calcium propionate, calcium succinate, caldaret, calusterone, camazepam,camostat, camphor, camphorate, camphotamide, camptothecin, candesartan,candesartan cilexetil, candoxatril, canertinib, canrenone, cantharidin,cantuzumab mertansine, capecitabine, capobenic acid, capravirine,capromab, capsaicin cream, captodiamine, captopril, captopril+HCTZ,capuride, carabersat, caramiphen, carazolol, carbachol, carbamazepine,carbamide peroxide, carbarsone, carbaryl, carbazochrome, carbendazim,carbenicillin, carbenoxolone, carbetapentane, carbicarb, carbidopa,carbidopa+levodopa-1, carbimazole, carbinoxamine, carbocloral,carbocysteine, carbon tetrachloride, carbonate gel, carboplatin,carboprost, carboprost, carboquone, carbromal, carbubarb, carbutamide,carbuterol, carfimate, carglumic acid, cargutocin, carindacillin,cariporide, cariporide, carisoprodol, carmofur, carmoxirole, carmustine,carnitine, caroverine, caroxazone, carphenazine, carpipramine,carprofen, carsalam, carteolol, carticaine, carubicin, carumonam,carvacrol, carvedilol, carvone, cascarillin, caspofungin, catechin,cathepsin K inhibitors, cathepsin S inhibitors, CC-401, CCI-779, CCR5antagonists, CDC-394, CDC-801, CEE-03-310, cefactor, cefadroxil,cefalexin, cefalexin pivoxil, cefamandole, cefatrizine, cefazedone,cefazolin, cefbuperazone, cefcapene pivoxil, cefclidin, cefdinir,cefditoren pivoxil, cefepime, cefetamet, cefetamet pivoxil, cefixime,cefinenoxime, cefmetazole, cefminox, cefodizime, cefonicid,cefoperazone, cefoperazone+sulbactam, ceforanide, cefoselis, cefotazime,cefotetan, cefotiam, cefotiam hexetil, cefoxitin, cefozopran,cefpimizole, cefpiramide, cefpirome, cefpodoxime, cefprozil,cefroxadine, cefsulodin, ceftazidime, cefteram, ceftezole, ceftibuten,ceftizoxime, ceftizoxime, ceftriaxone, cefuroxime, cefuroxime axetil,cefuzonam, celecoxib, celgosivir, celiprolol, cellulose ethyl, CEP-1347,CEP-701, cephacetrile, cephaeline, cephalexin, cephaloglycin,cephaloridine, cephalosporin C, cephalothin, cephapirin, cephradine,cerivastatin, ceronapril, certoparin, ceruletide, cerviprost,cetalkonium, cetamolol, cethexonium, cethromycin, cetiedil, cetirizine,cetirizine, cetirizine+pseudoephedrine, cetotiamine, cetoxime,cetraxate, cetrimonium, cetrorelix, cetyldimethylethyl-ammonium,cetylpyridinium, cevimeline, CG-1521, chaulmoogric acid, chenodiol,CHF-3381, chlophedianol, chloracizine, chloral, chlorambucil,chloramine-B, chloramine-T, chloramino-chloramphenicol, chlorazanil,chlorbenzoxamine, chlorbetamide, chlorcyclizine, chlordantoin,chlorguanide, chlorhexadol, chlorhexidine, chloriazepoxide,chlorisondamine, chlormadinone, chlormerodrin, chlormezanone,chlormidazole, chlornaphazine, chloroazodin, chlorophyll,chloroprednisone, chloroprocaine, chloropyramine, chloroquine,chlorothen, chlorothiazide, chlorotrianisene, chloroxine, chloroxylenol,chlorozotocin, chlorphenamine, chlorphenesin, chlorpheniramine,chlorphenoxamide, chlorphenoxamine, chlorphentermine, chlorproethazine,chlorproguanil, chlorproguanil+dapsone, chlorpromazine, chlorpropamide,chlorprothixene, chlorquinaldol, chlortetracycline, chlorthalidone,chlorthenoxazine(e), chlorzoxazone, cholic acid, choline, cholinetheophyllinate, choline-L-alfoscerate, chromocarb, chromonar,chrysoidine, CHS-828, CI-1031, CI-1040, cibenzoline, ciclesonide,cicletanine, ciclonicate, ciclopirox, ciclosidomine, ciclosporin A,cidofovir, cifenline, cilansetron, cilastatin, cilazapril, cilengitide,cilnidipine, cilomilast, cilostazol, cimetidine, cimetropium,cinacalcet, cinchonidine, cinchonine, cinchophen, cinepazet, cinepazide,cinepazide, cinitapride, cinmetacin, cinnamedrine, cinnarizine,cinolazepam, cinoxacin, cinoxate, cinromide, cioteronel, cipamfylline,cipralisant, ciprofibrate, ciprofloxacin, ciprofloxacin+ciramadol,cisapride, cisatracurium, cisplatin, citalopram, citicoline, Citiolone,citrate, citric acid, citrulline, cizolirtine, CJ-13610, CKD-602,cladribine, clanobutin, clarithromycin, clavulan, clavulanate disodium,clavulanic acid, clebopride, clemastine, clemizol, clenbuterol,clentiazem, clevidipine, clevudine, clidanac, clidinium, clinafloxacin,clindamycin, clindamycin, clindamycin+tretinoin, clinofibrate,clinprost, clobazam, clobenfurol, clobenoside, clobenzepam, clobenzorex,clobenztropine, clobetasol, clobetasone, clobutinol, clocapramine,clocinizine, cloconazole, clocortolone, clodronate, clodronic acid,clofarabine, clofazimine, clofenamide, clofibrat, clofibric acid,cloflucarban, clofoctol, cloforex, clomacran, clomestrone, clometacin,clomethiazole, clometocillin, clomiphene, clomipramine, clomocycline,clonazepam, clonidine, clonitazene, clonitrate, clonixin, clopamid,clopenthixol, cloperastine, clopidogrel, clopirac, cloprednol,cloranolol, clorazepic acid, clorexolone, cloricromene, clorindione,clorprenaline, clortermine, clospirazine, clostebol, clothiapine,clotiazepam, clotrimazole, clotrimazole+betamethasone, cloxacillin,cloxazolam, cloxotestosterone, cloxyquin, clozapine, CMI-392, CMT-3,CNI-1493, CNS-5161, cobamamide, cocaethylene, cocaine, codeine,cofactor, colchicine, colesevelam, colestilan, colestipol, colforsindaropate, colfosceril, collagraft, colocynthin, colpormon, coluracetam,combretastatin A-4 prodrug, compound B, conivaptin conjugate,connettivina, convallatoxin, coparaffinate, corticorelin ovine,corticosterone, cortisone, cortivazol, cosyntropin, cotarnine, cotinine,co-trimazine, coumetarol, CP-248, CP-461, CPC-211, CPI-1189, CRA-0450,creatinol-O-phosphate, CRL-5861, crobenetine, croconazole, cromoglicicacid, cromolyn, cropropamide, crotamiton, crotethamide, crystacide,CS-502, CS-758, CS-834, CT-052923, CT-32228, cupric citrate,cuproxoline, CVT-2584, CX-659S, cyacetacide, cyamemazine, cyanidin,CYC400, cyclacillin, cyclandelate, cyclazocine, cyclexanone,cyclexedrine, cyclidrol, cyclin D1 inhibitors, cyclizine, cyclobarbital,cyclobendazole, cyclobenzaprine, cyclobutyrol, cyclocumarol, cyclodrine,cyclofenil, cycloguanil, cyclomethycaine, cycloniumelodide,cyclopentamine, cyclopenthiazide, cyclopentobarbital, cyclopentolate,cyclophosphamide, cyclopiroxalamine, cycloserine, cyclothiazide,cyclovalone, cymarin, cymserine, cynarin(e), cyp26 inhibitors,cyproheptadine, cyproterone, cysteamine, cystic fibrosis ther,cytarabine, D-24851, D-4418, DA-5018, DA-6034, DA-7867, DA-7911,DA-8159, dacarbazine, daclizumab, dactinomycin, dalbavancin,dalfopristin, dalfopristin+quinupristin, dalteparin, daltroban,danaparoid, danazol, danthron, dantrolene, dapiprazole, dapivirine,dapoxetine, dapsone, daptomycin, darbepoetin alfa, darifenacin,daunorubicin, DAX<SciClone, DB-67, D-camphocarboxylic, DCF-987, DDT,deaminooxytocin, deanol, debrisoquin, decamethonium, decimemide,decitabine, declopramide, deferiprone, deferoxamine, deflazacort,defosfamide, degarelix, dehydroascorbic acid, dehydroemetine,dehyrdocholic acid, delapri+manidipine, delapril, delavirdine,delmadinone, delmopinol, delorazepam, delucemine, demanyl, demecarium,demeclocycline, demecolcine, demegestone, demexiptilline, denaverine,dendrimers, denileukin diftitox, denopamine, denopterin, deoxycholicacid, deoxycorticosterone, deoxydihydro-streptomycin, deoxyepinephrine,depreotide, depsipeptide, deptropine, dequalinium, dersalazine,deserpidine, desferrioxamine, desflurane, desipramine, deslanoside,desloratadine, deslorelin, desmopressin, desogestrel,desogestrel+estradiol, desogestrel+ethinylestrad (1), desomorphine,desonide, desoximetasone, detaxtran, devacade, dexamethasone,dexanabinol, dexecadotril, dexefaroxan, dexetimide, dexibuprofen,dexketoprofen, dexloxiglumide, dexmedetomidine, dexmethylphenidate,dexpanthenol, dexrazoxane, dextran-1, dextranomer, dextroamphetamine,dextromethorphan, dextromoramide, dextropropoxyphene, dezocine, DF-1012,DFA-IV, D-fenchone, D-glucuronolactone, Diab II, diacerein, diampromide,diamthazole, diathymosulfone, diatrizoate, diazepam, diaziquone,diazoxide, dibekacin, dibenzepin, dibromopropamidine, dibucaine,dichloralphenazone, dichloramine T, dichlorisone, dichlorobenzylalcohol, dichlorohydrin, dichlorophen, dichlorophenarsine,dichlorphenamide, diclofenac, diclofenac+HA, dicloxacillin, dicoumarol,dicumarol, dicyclomine, didanosine, dideoxyadenosine, didox, dienestrol,dienogest, dienogest+estradiol, diethadione, diethazine, diethylamide,diethylbromo-acetamide, diethylcarbamazine, diethylpropion,diethylstilbestrol, difemerine, difenamizole, difenoxin, difenpiramide,diflomotecan, diflorasone, difloxacin, diflucortolone, diflunisal,difluprednate, digitalin, digitoxin, digoxin, dihexyverine,dihydralazine, dihydrocodeine, dihydrocodeinone enol,dihydroergocryptine, dihydroergocryptine, dihydroergotamine,dihydromorphine, dihydrostreptomycin, dihydrotachysterol,dihydroxyaluminum, diisopromine, diisopropyl paraoxon, diisopropylamine,dilazep, dilevalol, diloxanide, diltiazem, dimecrotic acid, dimefline,dimeglumine, dimemorfan, dimenhydrinate, dimenoxadol, dimepheptanol,dimercaprol, dimetacrine, dimethadione, dimethazan, dimethindene,dimethisoquin, dimethisterone, dimethocaine, dimethoxanate, dimethylsulfoxide, dimethylthiambutene, dimetofrine, dimorpholamine,dinoprostone, diosmectite, diosmin, dioxadrol, dioxaphetyl,dioxethedrine, dioxybenzone, diphemanil, diphenadione, diphencyprone,diphenhydramine, diphenidol, diphenoxylate, diphenylpyraline,diphetarsone, diphtheria & tetanus toxoids and acellular pertussisvaccine adsorbed, dipipanone, dipivefrin, dipyridamole, dipyridamole,dipyrocetyl, dipyrone, diquafosol, dirithromycin, disodium pamidronate,disofenin, disopyramide, distigmine, disulfamide, disulfiram, ditazol,dithiazanine, dithranol, ditiocarb, dixanthogen, dixyrazine, DJ-927,DK-507k, DL-Lactic Acid, DMDC, DMXAA, DNA Stealth, dobesilate,dobutamine, docarpamine, docetaxel, docosahexaenoic acid, docosanol,docusate, dofetilide, dolasetron mesilate, domiodol, domiphen,domitroban, domperidone, donepezil, donitriptan, dopamine, dopexamine,doramapimod, doranidazole, doripenem, dorzolamide, dorzolamide+timolol,dosmalfate, dosulepine, dotarizine, dothiepin, doxacurium, doxapram,doxazosin, doxefazepam, doxenitoin, doxepin, doxercalciferol,doxifluridine, doxofylline, doxorubicin, doxycycline, doxylamine,DPC-817, DPI-3290, DQ-113, drofenine, droloxifene, drometrizole,dromostanolone, dronabinol, dronedarone, droperidol, droprenilamine,dropropizine, drospirenone, drotaverine, drotebanol, droxicam,droxidopa, droxidopa, DU-125530, duloxetine, duramycin, durapatite,dutasteride, DW-1141, DW-286a, DW-471, DX-9065a, DY-9760e, dyclonine,dydrogesterone, dymanthine, dyphyllin, E-1010, E-2101, E2F antagonists,E-3620, E-5564, E-5842, E-6259, EAA-90, ebastine, eberconazole,ebrotidine, ebselen, eburnamonine, ecabapide, ecabet, ecadotril,ecgonidine, ecgonine, echothiophate, econazole, ecopipam, ecraprost,ectylurea, ED-71, edaravone, edatrexate, edetate calcium disodium,edetate disodium, edetate sodium, edetate trisodium, edonentan,edotreotide, edoxudine, edrecolomab, edrophonium, efalith, efaproxiral,efavirenz, efletirizine, eflornithine, efloxate, eflucimibe,efonidipine, EGIS-7229, eglumegad, egualen, elarofiban, elcatonin,elcosapentaenoic acid, eledoisin, eletriptan, elgodipine, ellagic acid,elliptinium, eltoprazine, elvucitabine, elzasonan, embelin, embramine,emedastine, emepronium, emetine, emitefur, EMM-210525, emodin,emorfazone, EMR-62203, emtricitabine, emylcamate, enalapril,enalaprilat, enallylpropymal, encainide, enciprazine, endralazine,enfenamic acid, enflurane, enilconazole, eniluracil, ENMD-0995,enocitabine, enol-3-IPA, enoxacin, enoxaparin, enoximone, enoxolone,enprostil, enrasentan, entacapone, entecavir, enviomycin, eoinephrine,epalrestat, epavir, EPC-K1, eperisone, epervudine, ephedrine, epicillin,epimestrol, epinastine, epirizole, epirubicin, epitiostanol, eplerenone,eplivanserin, epoprostenol, epostane, eprazinone, epristeride,eprosartan, eprozinol, eptapirone, eptaplatin, eptastigmine, eptazocine,eptifibatide, equilenin, equilin, ERA-923, erdosteine, ergocornine,ergocorninine, ergoloid mesylates, ergonovine, ergosterol, ergotamine,eritadenine, erlotinib, ertapenem, erythrityl tetranitrate,erythrocentaurin, erythromycin acistrate, erythromycin erythrophleine,erythromycin estolate, erythromycin glucoheptonate, erythromycinlactobionate, erythromycin propionate, erythromycin stearate,erythromycin stinoprate, esaprazole, escitalopram, esculin, eseridine,esmolol, esomeprazole, estazolam, ester, estradiol, estradiol,estramustine, estriol, estrogen, estrone, eszopiclone, etafedrine,etafenone, etamiphyllin, etanercept, etanidazole, etaqualone, eterobarb,ethacridine, ethacrynic acid, ethadion, ethambutol, ethamivan,ethamsylate, ethanolamine, ethaverine, ethchlorvynol, ethenzamide,ethiazide, ethinamate, ethinyl estradiol, ethinyl estradiol, ethinylestradiol, ethionamide, ethisterone, ethoheptazine, ethopropazine,ethosuximide, ethotoin, ethoxzolamide, ethybenztropine, ethyl alcohol,ethyl biscoumacetate, ethyl chloride, ethyl dibunate, ethyl ether, ethylicosapentate, ethyl loflazepate, ethyl loflazepate, ethylamine,ethylene, ethylestrenol, ethylidene, ethylmethyl-thiambutene,ethylmorphine, ethylnorepinephrine, ethynodiol, ethynylcytidine,etidocaine, etidronate, etidronic acid, etifelmin, etifoxine, etilefrin,etilevodopa, etiprednol, etiroxate, etizolam, etodolac, etodroxizine,etofenamate, etofibrate, etofylline, etofylline clofibrate, etofyllinenicotinate, etoglucid, etomidate, etomidoline, etonitazene,etonogestrel, etoperidone, etoposide, etoposide phosphate, etoricoxib,etoxadrol, etozolin, etretinate, etryptamine, etymemazine, eucatropine,eugenol, EUK-134, EUK-189, evans blue, everolimus, exalamide,exametazime, exatecan, exemestane, exifone, exisulind, Exosurf®,ezetimibe, Factor IX, Factor VIII, Factor XIII, fadolmidine, fadrozole,falecalcitriol, famciclovir, famotidine, fampridine, fandofloxacin,fantofarone, faropenem, faropenem daloxate, fasidotril, fasudil,fazadinium bromide, febarbamate, febuprol, febuxostat, fedotozine,felbamate, felbinac, felodipine, felypressin, femoxetine, fenbenicillin,fenbufen, fenbutrazate, fencamfamine, fencamine, fenclozic acid,fendiline, fendosal, fenethylline, fenfluramine, fenipentol,fenofibrate, fenoldopam, fenoprofen, fenoterol, fenoverine,fenoxazoline, fenoxedil, fenozolone, fenpentadiol, fenpiprane,fenpiverinium, fenproporex, fenquizone, fenretinide, fenspiride,fentanyl, fentiazac, fenticlor, fenticonazole, fentonium bromide,fepradinol, feprazone, ferric sodium edetate, ferrioxamine B,ferrocholinate, ferrous gluconate, ferumoxytol, fesoterodine,fexofenadine, fibrostat, fidarestat, fiduxosin, finasteride, finrozole,fipexide, FK-960, flavopiridol, flavoxate, flecainide, fleroxacin,flesinoxan, flibanserin, floctafenine, flomoxef, flopropione,florantyrone, flosequinan, floxacillin, floxuridine, fluacizine,fluanisone, fluarizine, fluasterone, fluazacort, flucloronide,flucloxacillin, fluconazole, flucytosine, fludarabine, fludeoxyglucoseF18, fludiazepam, fludrocortisone, flufenamic acid, fluindione,flumazenil, flumecinol, flumequine, flumethasone, flumethiazide,flunisolide, flunitrazepam, flunoxaprofen, fluocinolone acetonide,fluocinolone SAL, fluocinonide, fluocortin butyl, fluocortolone,fluorescein, fluoresone, fluorometholone, fluorosalan, fluorouracil,fluoxetine, fluoxymesterone, flupentixol, fluperolone, fluphenazine,flupirtine, fluprednidene acetate, fluprednisolone, fluproquazone,flurandrenolide, flurazepam, flurbiprofen, flurithromycin, flurogestone,flurothyl, fluroxen, fluspirilene, flutamide, flutazolam, fluticasone,flutoprazepam, flutrimazole, flutropium bromide, fluvastatin,fluvoxamine, folic acid, folinic acid, fomepizole, fominoben,fomivirsen, fomocaine, fonazine, fondaparinux, formebolone, formestane,formocortal, formoterol, fosamprenavir, foscarnet, fosfestrol,fosfluconazole, fosfomycin, fosfomycin, fosfosal, fosinopril,fosphenytoin, fotemustine, fropenem, frovatriptan, fructose,fructose-1,6-diphosphate, FTC, FTY-720, fudosteine, fulvestrant,fumagiline, fumagillin, furaltadone, furazabol, furazolidone, furazoliumchloride, furonazide, furosemide, fursultiamine, furtrethonium, fusidicacid, GI, YM BioSciences, G25, GABA-A Alpha5, gabapentin, gabexate,gaboxadol, gadobenat, gadobutrol, gadodiamide, gadolinium, gadopenteticacid, gadoteridol, gadoversetamide, gadoxetic acid, galantamine,galanthamine, galarubicin, gallamine triethiodide, gallic acid, galliummaltolate, gallium nitrate, gallopamil, ganaxolone, ganciclovir,ganirelix, ganstigmine, gantofiban, garenoxacin, garnocestim,gatifloxacin, gefarnate, gefitinib, gemcabene, gemcitabine, gemeprost,gemfibrozil, gemifloxacin, gentamicin, gentian violet, gentiopicrin,gentisic acid, gepefrine, gepirone, gestodene, gestodene+ethinylest,gestonorone caproate, gestrinone, gimatecan, giractide, gitoxin,GL-406349, Glafenine, glatiramer, Glibornuride, gliclazide, glimepiride,glipizide, gliquidone, glisolamide, glisoxepid, globulin (human),glucametacin, glucoheptonic acid, gluconic acid, glucosamine,glucosulfone, glufosfamide, glutamic acid, glutaraldehyde, glutethimide,glyburide, glybuthiazol(e), glybuzole, glycerol, glycerophosphate,glycocyamine, glycol salicylate, glyconiazide, glycopyrrolate,glyhexamide, glymidine, glypinamide, GMDP, gold sodium, goserelin,GPI-1485, GPI-5693, graftskin, granisetron, grepafloxacin, griseofulvin,guaiacol, guaiapate, guaiazulene, guaifenesin, guaimesal,gualacolsulfonate, guamecycline, guanabenz, guanadrel, guanethidine,guanfacine, guanoxabenz, guanoxan, gugulipid, gusperimus, GW-280430A,GW-320659, GYKI-16084, hachimycin, halazepam, halcinonide, halobetasol,halofantrine, halometasone, haloperidol, halopredone, haloprogin,halopropane, halothane, haloxazolam, harkoseride, HE-2000, healos,hematoporphyrin, hepronicate, heptabarbital, heptaminol, hetacillin,hetastarch, hexacetonide, hexachlorophene, hexadimethrine,hexafluorenium, hexamethonium, hexamidine, hexapropymate, hexedine,hexestrol, hexestrol Bis(β-diethylaminoethyl ether), hexethal,hexetidine, hexobarbital, hexobendine, hexocyclium methyl sulfate,hexoprenaline, hextend, hexylcaine, HF-0299, HGP-2, HGP-6A, hidrosmin,histamine, Histapyrrodine, histrelin, HM-101, HMN-214, homatropine,homocamfin, homochlorcyclizine, hopantenic acid, HP-228, huperzine A,hyaluronan, hycanthone, hydnocarpic acid, hydralazine, hydrastine,hydrastinine, hydrochlorothiazide, hydrocodone, hydrocortamate,hydrocortisone, hydrocortisone, hydroflumethiazide, hydromorphone,hydroquinidine, hydroquinine, hydroquinone, hydroxid, hydroxocobalamin,hydroxyamphetamine, hydroxychloroquine, hydroxydione, hydroxyethylether, hydroxynaphthoate, hydroxypethidine, hydroxyphenamate,hydroxypropyl cellulose, hydroxystilbamidine, hydroxytetracaine,hydroxyzine, Hylan G-F 20, hymecromone, hyoscyamine, hypericin, IACFT,ibandronic acid, ibopamine, ibopamine, Ibritumomab, ibrolipim,ibudilast, Ibufenac, ibuprofen, ibuprofen piconol, ibuproxam, ibutilide,ICA-17043, icodextrin, idarubicin, Idazoxan, IdB-1016, idebenone,IDN-5109, idoxifen, idraparinux, idrocilamide, ifenprodil, ifosfamide,iguratimod, ilaprazole, ilomastat, iloperidone, iloprost trometamol,ILX23-7553, imatinib, imidapril, imidazole salicylate, imipenem,imipramine, imipramine N-Oxide, imiquimod, imolamine, implitapide,improsulfan, inactivated, inaperisone, incadronate, incadronic acid,indalpine, indanazoline, indapamide, indecainid, indeloxazine,indeloxazine, indenolol, indinavir, indiplon, indisetron, indisulam,indobufen, indocyanine green, indometacin, indoprofen, indoramin,induclem, infliximab, inhibitor, inhibitors, inosine pranobex, inositol,inositol niacinate, inverse agonist Mer, iobenguane, iobenzamic acid,iobitridol, iocarmic acid, iocetamic acid, iodamide, iodide, iodine,iodipamide, iodixanol, iodoalphionic acid, iodochlorhydroxyquin,iodoform, iodopyracet, iodopyrrole, iodoquinol, iodosubgallate,iofetamine 123I, ioglycamic acid, iohexol, iomeglamic acid, iomeprol,iopamidol, iopanoic acid, iopentol, iophendylate, iophenoxic acid,iopromide, iopronic acid, iopydol, iopydone, iothalamic acid, iotrolan,ioversol, ioxaglic acid, ioxilan, IP-751, ipidacrine, IPL-576092,ipodate, iponiazid, ipratpopium, ipratropium, ipratropium bromide,iprazochrome, ipriflavone, iprindole, iproclozid, ipsapiron, irbesartan,IRFI-042, IRFI-165, iridomyrmecin, irindalone, irinotecan, irofulven,iron sorbitex, irsogladine, IS-741, isaglitazone, ISAtx-247, isbogrel,isepamicin, isoaminile, isobutyl p-aminobenzoate, isoconazole,isoetharine, isofloxythepin, isoflurane, isoflurophate, isoladol,isomethadone, isometheptene, isoniazid, isonixin, isopromethazine,isopropamide iodide, isopropyl alcohol, isopropyl unoprostone,isoproterenol, isosorbide, isosorbide dinitrate, isosorbide mononitrate,isothipendyl, isotretinoin, isovaleryl, isoxepac, isoxicam, isoxsuprine,isradipine, israpafant, ISV-403, itasetron, ITF-282, itopride,itraconazole, itramin, itriglumide, iturelix, ivabradine, ixabepilone,J-104132, J-107088, J-113397, Janex-1, josamycin, JTV-519, K-777, kainicacid, kalimate, kallidin, KB-130015, KCB-328, kebuzone, ketamine,ketanserin, ketazolam, kethoxal, ketobemidone, ketoconazole, ketoprofen,ketorolac, ketorolac, ketotifen, khellin, kinetin, KNI-272, KP-103,KP-157, KP-544, KRN-5500, KT-136, KUL-7211, KW-2170, KW-6002, KW-7158,L-365260, L-5-hydroxy-tryptophan, L-745337, L-758298, L-826141,labetalol, lacidipine, lactic acid, lactitol, lactulose, lafutidine,lamifiban, lamivudine, lamotrigine, landiolol, lanicemine, laniquidar,lanoconazole, lanoteplase, lanreotide, lansoprazole, lanthanumcarbonate, lapatinib, laquinimod, lasofoxifene, latamoxef, latanoprost,lauroguadine, laurolinium acetate, lawsone, LAX-111, lazabemide,LB-30057, L-cysteine, lefetamine, leflunomide, leflunomide, leiopyrrole,lenampicillin, lentinan, lepirudin, lercanidipine, lerisetron,lesopitron, leteprinim, letosteine, letrozole, leucocyanidin,leuprolide, leuprolide acetate, leuprorelin, levallorphan, levaminsole,levcromakalim, levetiracetam, levobetaxolol, levobunolol,levobupivacaine, levocabastine, levocetirizine, levodopa,levodropropizine, levofloxacin, levomethadyl acetate, levomoprolol,levonorgestrel, levophacetoperane, levopropoxyphene, levorphanol,levosimendan, levosulpride, levothyroxine, levovirin, lexidronam,lexipafant, LF-15-0195, LF-16-0687, LGD-1550, LH, LH-RH, liarozote,licofelone, licostinel, lidadronate, lidamidine, lidocaine, lidofenin,lidoflazine, limaprost, lincomycin, lindan, linezolid, linoleic acid,linolenic acid, liothyronine, lipase, lipo-dexamethasone,lipo-flurbiprofen, Lipogel HA, LiquiVent, liranaftate, lisinopril,lisofyllin, lisuride, lithium, lithium citrate, lixivaptan, LJP-1082,LLUAlpha, LMP-160, LMP-420, loanzapine, lobaplatin, lobeline,lobenzarit, lodoxamide, lofentanil, lofepramine, lofexidine,loflucarban, lomefloxacin, lomerizine, lomifylline, lomustine,lonafarnib, lonapalene, lonazolac, lonidamine, loperamide, loperamideoxide, loprazolam, loprinone, loracarbef, lorajmine, loratadine,lorazepam, lorcainide, lormetazepam, lornoxicam, losartan, loteprednol,lotrafiban, lovastatin, loxapine, loxiglumide, loxoprofen, Lu-35-138,lubeluzole, lubiprostone, lucanthone, lucanthone, lumefantrine,lumiracoxib, lurtotecan, lutetium texaphyrin, LV-216, LX-104, LY-156735,LY-293111, LY-293558, LY-355703, lyapolate, lymecycline, lynestrenol,lypressin, lysine acetylsalicylate, lysine salicylate,lysophospholipids, M-40403, mabuprofen, mabuterol, macrophagecolony-stimulating factor, MADU, mafenide, mafosfamide, magaldrate,magenta I, magnesium, magnesium carbonate, magnesium chloride, magnesiumcitrate, magnesium gluconate, magnesium lactate, magnesium salicylate,malathion, malotilate, mandelic acid, mandelic acid isoamyl,mangafodipir, manidipine, mannomustine, mannose-6-phosphate,maprotilline, maribavir, marimastat, maxacalcitol, mazindol,mazipredone, MC-5723, MCC-478, MCI-154, m-cresyl acetate, MDAM, MDI-101,MDI-403, MDL-100907, mebendazole, mebeverine, mebhydroline, mebrofenin,mebutamate, mecamylamine, mechlorethamine, mechlorethamine oxide,mecillinam, meclizine, meclocycline, meclofenamate, meclofenamic acid,meclofenoxate, mecloqualone, mecysteine, medazepam, medifoxamine,medrogestone, medronic acid, medroxyprogesterone, medrysone, mefenamicacid, mefenorex, mefexamide, mefloquine, mefruside, megestrol, meglumin,meglutol, melagatran, melanocortin-4 agonist, melarsoprol, melengestrol,melevodopa, melinamide, melitracen, meloxicam, melperone, melphalan,meluadrine, memantine, MEN-10700, MEN-10755, menadiol, menadione,menadoxime, menbutone, menogaril, MENT, menthol, menthyl valerate,meobentine, meparfynol, mepartricin, mepazine, mepenzolate bromide,meperidine, mephenesin, mephenoxalone, mephentermine, mephenytoin,mephobarbital, mepindolol, mepitiostane, mepivacaine, mepixanox,meprednisone, meprobamate, meproscillarin, meptazinol, mequitazine,meralein, meralluride, merbromin, mercaptomerin, mercumallylic acid,mercuric oleate, mercuric oxycyanide, merimepodib, meropenem, mersalyl,mertiatide, mesalamine, mesalazine, mesna, mesoridazine, mestanolone,mesterolone, mestranol, mesulfen, metaclazepam, metampicillin,metapramine, metaproterenol, metaraminol, metazocine, metergoline,metformin, methacholine, methacycline, methadone, methafurylene,methamphetamine, methandriol, methandrostenolone, methantheline,methapyrilene, methaqualone, metharbital, methazolamide, methdilazine,methenamine, methenolone, methestrol, methetoin, methicillin,methimazole, methiodal, methionic acid, methionine, methisazone,methitural, methixene, methocarbamol, methohexital, methotrexate,methotrimeprazine, methoxamine, methoxsalen, methoxycinnamate,methoxyflurane, methoxyphenamine, methoxypromazine, methscopolamine,methsuximide, methyclothiazide, methyl blue, methyl nicotinate, methylpropyl ether, methyl salicylate, methyl tert-butyl ether,methylbenzethonium chloride, methylbromide, methylcobalamin, methyldopa,methylene blue, methylergonovine, methylhexaneamide, methylphenidate,methylprednisolone, methylprednisolone, methylprednisolone,methylthiouracil, methyltrienolone, methyprylon, methysergide,metiazinic acid, metipranolol, metoclopramide, metocurine iodide,metofenazate, metolazone, metopimazine, metopon, metoprolol,metralindole, metrizamide, metrizoic acid, metron s, metyrapone,metyrosine, mexazolam, mexenone, mexiletine, mezlocillin, MFH-244,mianserin, mibefradil, miboplatin, micafungin, miconazole, micronomicin,midaxifyline, midazolam, midecamycin, midecamycin acetate, midesteine,midodrine, midostaurin, mifepristone, miglitol, miglustat, mildronate,milnacipran, miloxacin, milrinone, miltefosine, minaprine, minocycline,minodronic acid, minoxidil, miokamycin, mirtazapine, misoprostol,mitemcinal, mitiglinide, mitobronitol, mitoguazone, mitolactol,mitomycin, mitotane, mitoxantrone, mitoxantrone, MIV-210, mivacurium,mivazerol, mizolastine, mizoribine, MKC-733, MLN-519, MLN-576,moclobemide, modafinil, moexipril, mofarotene, mofebutazone, mofegiline,mofetil, mofezolac, MOL-6131, molindone, molsidomine, mometasone,monatepil, monobenzone, monoethanolamine, monolaurin, monoterpene diols,montelukast, monteplase, moperone, mopidamol, moprolol, moracizine,morazone, moricizine, moroxydine, morphazinamide, morphine,morphine-6-glucuronide, mosapramine, mosapride, motexafin, motretinide,moveltipril, moxalactam, moxastine, moxaverine, moxestrol, moxifloxacin,moxisylyte, moxonidine, M-PGA, MPI-5010, MPI-5020, MPL, MRS-1754,MS-209, MS-275, MS-325, MS-377, mupirocin, muscarin, muzolimine,MX-1013, mycophenolate, mycophenolic acid, myrophine,N-(hydroxymethyl)-nicotinamide, N,N,N′,N′-tetraethylphthalamide,N-[4-[4-(2-methoxyphenyl)-1-piperazinyl]butyl]naphthalene-2-carboxamide,N2-formyl-sulfisomidine, N4-sulfanilylsulfanilamide,N4-β-D-glucosylsulfanilamide, nabilone, nabumetone, N-acetylcysteine,N-acetylmethionine, nadifloxacin, nadolol, nadoxolol, nafamostat,nafarelin, nafcillin, nafronyl, naftidofuryl, naftifine, naftopidil,nalbuphine, nalidixic acid, nalmefene, nalorphine, naloxone, naltrexone,NAMI, naminidil, nandrolone, napadisilate, naphazoline, naphthalene,naproxen, naproxen betainate, naratriptan, narceine, narcobarbital,natamycin, nateglinide, N-butyldeoxy-nojirimycin, N-butylscopolammoniumBromide, NC-503, NC-531, NCX-1000, NCX-4016, NCX-456, NCX-950,n-docosanol, NE-100, nealbarbital, nebivolol, nebostinel, nebracetam,nedaplatin, nedocromil, nefazodone, nefiracetam, nefopam, negamycin,nelfinavir, nemonapride, neostigmine, nepadutant, neramexane, neridronicacid, neriifolin, N-ethylamphetamine, neticonazole, netilmicin,nevirapine, NGD-98-2, nialamide, niaprazine, nicametate, nicaraven,nicardipine, nicergoline, niceritrol, niclosamide, nicoclonate,nicofuranose, nicomol, nicomorphine, nicorandil, nicotinamide, nicotine,nicotinic acid, nicotinic acid benzyl ester, nicotinyl alcohol,nifedipine, nifekalant, nifenalol, niflumic acid, nifuratel,nifurfoline, nifuroxazide, nifuroxime, nifurpirinol, nifurprazine,nifurtimox, nifurtoinol, nifurzide, NIK-254, nikethamide, nilutamide,nilvadipine, nimesulide, nimetazepam, nimodipine, nimorazole, nimustine,ninopterin, NIP-142, NIP-531, niperotidine, nipradilol, niridazole,nisoldipine, nitazoxanide, nitisinone, nitracrine, nitrazepam,nitrendipine, nitroflurbiprofen, nitrofurantoin, nitrofurazone,nitroglycerin, nitromersol, nitronaproxen, nitroxazepine, nitroxoline,nizatidine, nizofenone, NM-3, NM-702, N-methylephedrine,N-methylepinephrine, N-methylglucamine, NN-414, NNC-05-1869, nobel,nogalamycin, nolatrexed, nolomirole, nolpitantium, nomegestrol,nomifensine, noprylsulfamide, norbolethone, nordazepam, nordefrin,nordihydroguaiaretic acid, norelgestromin, norepinephrine,norethandrolone, norethindrone, norethynodrel, norfenefrine,norfloxacin, norgesterone, norgestimate, norgestrel, norgestrienone,norlevorphanol, normethadone, normethandrone, normorphine, norphenazone,norpipanone, norpseudoephedrine, nortriptyline, norvinisterone,noscapine, novembichin, novobiocin, noxiptillin, noxythiolin, NS-1209,NS-1231, NS-126, NS-220, NS-2330, NS5A inhibitors, NS-7, NS-8,NSC-330507, NSC-619534, NSC-697726, N-sulfanilyl-3,4-xylamide, NU-6027nucleosides, NV-07, NVP-SRA880, NW-1029, NXY-059, Nylidrin, NZ-314,NZ-419, obidoxime chloride, OC-108, ocinaplon, octabenzone, octacaine,octamoxin, octaverine, octenidine, octodrine, octopamine, octotiamine,octreotide, octyl, ofloxacin, oleandrin, oleic acid,olmesartan-medoxomil, o-lodohippurate, olopatadine, olpadronic acid,olsalazine, oltipraz, OM-294DP, omacor, omapatrilat, omeprazole,omiloxetine, omoconazole, onapristone, ondansetron, ONO-3403, ONO-4128,ONO-8815 Ly, ONT-093, OPC-14523, OPC-31260, OPC-51803, OPC-6535,opiniazide, opioid analgesics, opipramol, orazamide, orazipone,Org-12962, Org-24448, oritavancin, orlistat, ormeloxifene, ornidazole,ornipressin, ornithine, ornoprostil, orotic acid, orphenadrine,orthocaine, osalmid, osanetant, osaterone, oseltamivir, OSI-7836,OSI-7904, ospemifene, otilonium bromide, ouabain, oxaceprol, oxacillin,oxaflozane, oxaliplatin, oxalyt-C, oxamarin, oxametacine, oxamniquine,oxandrolone, oxantel, oxapropanium, oxaprozin, oxatomide, oxazepam,oxazolam, oxcarbazepine, oxeladin, oxendolone, oxethazaine, oxetoron,oxiconazole, oxidronic acid, oxiniacic acid, oxiracetam, oxitropium,oxolamin, oxolinix acid, oxophenarsine, oxprenolol, oxybenzone,oxybutynin, oxycinchophen, oxycodone, oxygent, oxymesterone,oxymetazoline, oxymetholone, oxymethurea, oxymorphone, oxypendyl,oxypertine, oxyphenbutazone, oxyphencyclimine, oxyphenisatin,oxyphenonium, oxypinocamphone, oxypurinol, oxytedrine, oxytetracycline,ozagrel, p-(benzylsulfonamido)-benzoic acid, P-100, P-1202, P32/98,PA-824, PACAP 38, pactitaxel, PADRE, pagoclone, PAI inhibs, palindore,palivizumab, palonosetron, pamabrom, pamaquine, pamicogral, pamidronate,p-aminobenzoic acid, p-aminohippuric acid, p-amino-propiophenone,p-aminosalicylic acid, panavir, pancuronium, panipenem, pantethine,pantoprazole, pantothenic acid, papain, papaverine, paracetamol,paraflutizide, paraldehyde, paramethadione, paramethasone, paranyline,parathyroid hormone, parecoxib, parethoxycaine, pargyline, paricalcitol,paromomycin, paroxetine, paroxypropione, parsalmide, patrin-2,pazinaclone, pazufloxacin, p-bromoacetanilide, PC-NSAIDs, PD-0166285,pecilocin, pefloxacin, pegvisomant, pelletierine, pemetrexed,pemirolast, pemoline, pempidine, PEN-203, penamecillin, penbutolol,penciclovir, penethamate, penfluridol, penicillamine, penicillin G,penicillin G Procaine, penicillin N, penicillin O, penicillin V,penimepicycline, penntuss, pentaerythritol, pentaerythritol,pentaerythritol chloral, pentagastrin, pentagestrone, pentalyte, pentamthonium, pentamidine, pentazocine, pentetate, pentetic acid,pentetreotide, penthienate, pentifyllin, pentigetide, pentisomide,pentobarbital, pentolinium, pentorex, pentosan, pentostatin,pentoxifylline, pentoxyl, pentrinitrol, pentylenetetrazole, peplomycin,peptide, peptide, perazine, perfiromycin, perflubron, perfosfamide,pergolide, perhexiline, pericyazine, perifosine, perillyl alcohol,perimethazine, perindopril, periodyl, perisoxal, perlapine,permanganate, permethrin, perospirone, perphenazine, petroleum benzin,PH-10, phanquinone, pharmacor, pharmaprojects no. 6362, pharmaprojectsno. 4994, pharmaprojects no. 5325, pharmaprojects no. 5972,pharmaprojects no. 6446, pharmaprojects no. 6590, pharmaprojects no.6656, pharmaprojects no. 6691, pharmaprojects no. 6743, pharmaprojectsno. 6748, phenacaine, phenacemide, phenacetin, phenadoxone, phenallymal,phenamet, phenamide, phenazocine, phenazopyridine, phenbutamide,phencyclidine, phendimetrazine, phenelzine, phenesterine,phenetharbital, phenethicillin, pheneturide, phenformin,phenglutarimide, phenindamine, phenindione, pheniprazine, pheniramine,phenmetrazine, phenobarbital, phenobutiodil, phenocoll, phenoctide,phenolphthalein, phenolphthalol, phenolsulfonphthalein,phenol-tetrachlorophthalein, phenoperidine, phenosulfazole,phenoxybenzamine, phenoxypropazine, phenprobamate, phenprocoumon,phenserine, phensuximide, phentermine, phentetiothalein, phentolamine,phenyl acetylsalicylate, phenyl aminosalicylate, phenyl salicylate,phenylbutazone, phenylephrine, phenylethanolamine, phenylmercury,phenylmethylbarbituric acid, phenylpropanolamine,phenylpropyl-methylamine, phenyltoloxamine, phenyramidol, phenytoin,phethenylate, phloroglucinol, pholcodine, pholedrine, phoramide,phosphate, phosphate, phosphocreatine, phosphocysteamine,phosphorylcholine, phthalylsulfathiazole, phthalysulfacetamide,p-hydroxyephedrine, phylloquinone, physostigmine, phytic acid, PI-88,piberaline, piboserod, picilorex, picloxydine, picoperine, picosulfate,picotamide, picumast, pidotimod, pifarnine, piketoprofen, pildralazine,pilocarpine, piloplex, pilsicainide, pimeclone, pimecrolimus,pimefylline, pimilprost, piminodine, pimobendan, pimozide, pinacidil,pinaverium, pinazepam, pindolol, pioglitazone, pipacycline, pipamazine,pipamperone, pipazethate, pipebuzone, pipecurium, pipecuronium,pipemidic acid, pipenzolate bromide, piperacetazine, piperacillin,piperazine adipate, piperidione, piperidolate, piperilate, piperineanalogues, piperocaine, piperonal, piperoxan, piperylone, pipobroman,piposulfan, pipotiazine, pipoxolan, pipradrol, piprozolin, piracetam,pirarubicin, pirazolac, pirbuterol, pirenoxine, pirenzepine, piretanide,pirfenidone, piribedil, piridocaine, pirifibrate, piritramide,piritrexim, pirlindole, pirmenol, piroctone, piroheptine, piromidicacid, piroxicam, piroxicam betadex, piroxicam cinnamate, pirozadil,pirprofen, pitavastatin, pivagabine, pivaloyloxymethyl,pivalylbenzhydrazine, pivampicillin, pivampicillin/pivmecillinam,pivcefalexin, pivmecillinam, pixantrone, pizotifen, pizotyline, PKI-166,p-lactophenetide, plafibride, plasminogen activator, plasmocid,platonin, plaunotol, PLD-118, PLD-147, pleconaril, plicamycin,p-methyldiphenhydramine, PMS-601, Pneumococcal, PNU-288034,podophyllotoxin, polaprezinc, poldine methylsulfate, policresulen,polidexide, polidocanol, poliovirus vaccine, poly-ADPRT inhibitors,polyestradiol, polyphenon E, polythiazide, porfimer, posaconazole,posatirelin, potassium, potassium, potassium, potassium chloride,potassium gluconate, potassium p-aminobenzoate, povidone,povidone-iodine, PP-17, PR-2699, PR-608, practolol, prajmaline,pralidoxime, pralnacasan, pramipexole, pramiracetam, pramiverin,pramlintide, pramoxine, pranidipine, pranlukast, pranoprofen,prasterone, pratosartan, pravastatin, prazepam, praziquantel, prazosin,prednicarbate, prednimustine, prednisolone, prednisolone21-diethylaminoacetate, prednisolone farnesil, prednisolone sodium,prednisone, prednival, prednylidene, pregabalin, pregnan-3α-ol-20-one,premarin+trimegestone, prenalterol, prenoxdiazine, prenylamine,prezatide, pridinol, prifinium, prilocaine, primaquine, primidone,prinomastat, PRO-2000, probenecid, probucol, procainamide, procaine,procarbazine, procaterol, prochlorperazine, procodazol, procyclidine,procymate, prodipine, proflavine, progabide, progesterone,proglumetacin, proglumide, proheptazine, prolactin, prolintane,prolonium, promazine, promedol, promegestone, promestriene,promethazine, pronethalol, propacetamol, propafenone, propagermanium,propallylonal, propamidine, propane-1,2-diol, propanidid, propantheline,proparacaine, propatyl, propenidazole, propentofylline, propicillin,propiomazine, propionic acid, propionyl 1-carnitine, propipocaine,propiram, propiverine, propizepine, propofol, propoxycaine,propoxyphene, propranolol, propylhexedrine, propyliodone,propylthiouracil, propyphenazone, proquazone, proscillaridin,prostacyclin, prostaglandin E1, prostaglandin E2, prostaglandin F2a,prosultiamine, protein C, protheobromine, prothipendyl, protiofate,protionamide, protizinic acid, protoanemonin, protoklol, protoporphyrinIX, protriptyline, pro-urokinase, proxazole, proxetil, proxibarbal,proxigermanium, proxyphylline, prozapine, prucalopride, prulifloxacin,pseudococaine, pseudoephedrine, pseudoephedrine,pseudoephedrine+triprolidine, psilocybin, PSK-3841,p-sulfanilyl-benzylamine, PT-141, pteropterin, puromycin, PX-12,pyrantel, pyrazinamide, pyridinol carbamate, pyridostigmine, pyridoxal5-phosphate, pyridoxine, pyrilamine, pyrimethamine, pyrinoline,pyrisuccideanol, pyrithione, pyrithyldione, pyritinol, pyrocatechol,pyrogallol, pyronaridine, pyrophosphate, pyrovalerone, pyroxylin,pyrrobutamine, pyrrocaine, pyrrolntrin, pyrvinium pamoate, quazepam,quercetin, quetiapine, quinacillin, quinacrine, quinagolide, quinapril,quinaprilat, quinapyramine, quinbolone, quinestradiol, quinestrol,quinethazone, quinfamide, quinidine, quinine, quinocide, quinupramine,quinupristin, R-107500, R-667, rabeprazole, racecadotril,racemethorphan, raloxifene, raltitrexed, ramatroban, ramifenazone,ramipril, ramosetron, Ramot project No. 1097, ranimustine, ranitidine,ranitidine bismuth, ranolazine, ranpirnase, rapacuronium, rasagiline,raubasine, ravuconazole, raxofelast, razoxane, RC-529, rebamipide,rebimastat, reboxetime, remacemide, remifentanil, reminetant,remoxipride, renzapride, repaglinide, repertaxin L-lysine salt,repinotan, repirinast, reposal, reproterol, rescimetol, rescinnamine,reserpiline, reserpine, resibufogenin, resiquimod, resorcinol,reteplase, retigabine, retinoic acid, revimid, R-flurbiprofen, rho (D)immune, rho-kinase inhibitors, ribavirin, riboflavin, ribostamycin,ricinoleic acid, ridogrel, rifabutin, rifalazil, rifametane, rifamide,rifampicin+trimethoprim, rifampin, rifamycin SV, rifapentine, rifaximin,rifaximine cream, rilmazafone, rilmenidine, riluzole, rimantadine,rimazolium, rimexolone, rimiterol, rimonabant, riodoxol, rioprostil,risedronate, risedronic acid, risperidone, ritanserin, ritipenem,ritodrine, ritonavir, rituximab, rivastigmine, rizatriptan, RJR-2403,RNA Stealth, Ro-0094889, Ro-61-1790, rociverine, rocuronium, rofecoxib,roflumilast, rokitamycin, rolipram, rolitetracycline, romurtide,ronifibrate, ropinirole, ropivacaine, roquinimex, rosaprostol,rosaramicin, rose bengal, rosiglitazone, rosoxacin, rostaporfin,rosuvastatin, rotigotine, rotraxate, roxarsone, roxatidine, roxifiban,roxindol, roxithromycin, RPR-109881A, RPR-130401, R-roscovitine,RS-0406, RSR-13, rubijervine, rubitecan, ruboxistaurin, rufinamide,rufloxacin, rupatadine, rutin, RWJ-54428, S-0139, S-15535, S-18886,S-34730, S-3578, S-36496, S-36527, S-5751, S-8510, S-8921, sabcomeline,sabeluzole, S-adenosylmethionine, safinamide, salacetamide,salazosulfadimidine, salbutamol, salicin, salicyl alcohol, salicylamide,salicylamide O-acetic acid, salicylanilide, salicylic acid,salicylsilfuric acid, salinazid, salmeterol, salsalate, salverine,samarium ¹⁵³Sm, sampatrilat, sancycline, saperconazole, sapropterin,saquinavir, saralasin, saredutant, saredutant, sarizotan, sarizotan,sarpogrelate, sarpogrelate, satigrel, satigrel, satraplatin,satraplatin, satumomab, satumomab, SB-237376, SB-237376, SB-238039,SB-238039, SB-277011, SB-277011, scarlet red, SCH-00013, SCH-00013,Sch-23863, Sch-23863, Sch-57790, Sch-63390, scillarenin, scopolamine,scopolamine, scopolamine N-oxide, SCS technology, secalciferol,secnidazole, secobarbital, selegiline, selenomethionine, sematilide,semotiadil, seocalcitol, sepimostat, seratrodast, sertaconazole,sertaconazole, sertindole, sertindole, sertraline, sertraline,sestamibi, setastine, setastine, sevelamer, sevelamer, sevoflurane,sevoflurane, SG-210, sibutramine, siccanin, sildenafil, silodosin,silprostone, silver lactate, silver picrate, silver sulfadiazine,simetride, simfibrate, simvastatin, sincalide, sintropium bromide,sisomicin, sitafloxacin, sitamaquine, sitaxsentan, sivelestat, SJA-6017,SL-65-1498, SLV-306, SLV-308, Sm153 lexidronam, S-methylmethionine,SMP-300, SN-38, SNAP-7941, SOA-132, soblidotin, sobrerol, sobuzoxane,sodium arsanilate, sodium arsphenamine, sodium chloride, sodiumdibunate, sodium folate, sodium formaldehydesulfoxylate, sodiumhyaluronate, sodium iodomethamate, sodium nitrite, sodium nitroprusside,sodium oxybate, sodium phenol-sulfonate, sodium phenylbutyrate, sodiumphosphate, sodium prasterone sulfate, sodium propionate, sodiumsalicylate, sodium tetradecyl sulfate, sofalcone, solasulfone,solifenacin, sorbinicate, sorbitol, sorivudine, sotalol, soterenol,sozoiodolic acid, spaglumic acid, sparfloxacin, sparteine, SPA-S-843,spasmolytol, SPD-754, spectinomycin, SPI-339, spiperone, spirapril,spirogermanium, spironolactone, SR-121463, SR-144190, SR-146131,SR-271425, SR-27897, SR-31747, SR-58611, SS732, SS-750, SSR-149415,SSR-180575, SSR-181507, SSR-591813, SST-101, SSY-726, ST-200,stachyfilin, stallimycin, stampidine, stannous, stannsoporfin,stanolone, stanozolol, staph aureus ther, STAT4 inhibitors, stavudine,stenbolone, stepronim, stibocaptate, stibophen, stilbamidine,stiripentol, streptodornase, streptomycin, streptonicozid,streptonigrin, streptozocin, strontium ranelate, strontium-89 chloride,succimer, succinimide, succinylcholine, succinylcholine,succinylsulfathiazole, succisulfone, suclofenide, sucralfate,sufentanil, sulbactam, sulbactam+ampicillin, sulbenicillin, sulbentine,sulbutiamine, sulconazole, suleptanate, sulesomab, sulfabenzamide,sulfacetamide, sulfachlorpyridazine, sulfachrysoidine, sulfacytine,sulfadiazine, sulfadicramide, sulfadimethoxine, sulfadoxine,sulfaethidole, sulfaguanidine, sulfaguanole, sulfalene, sulfaloxic acid,sulfamerazine, sulfameter, sulfamethazine, sulfamethizole,sulfamethomidine, sulfamethoxazole, sulfamethoxypyrazine,sulfamethoxypyridazine, sulfametrole, sulfamidochrysoidine, sulfamoxole,sulfanilamide, sulfanilic acid, sulfanilylurea, sulfaperine,sulfaphenazole, sulfaproxyline, sulfapyrazine, sulfapyridine,sulfarside, sulfarsphenamine, sulfasalazine, sulfasomizole,sulfasymazine, sulfathiazole, sulfathiourea, sulfinalol, sulfinpyrazone,sulfiram, sulfisomidine, sulfisoxazole, sulfobromophthalein,sulfonethylmethane, sulfoniazide, sulfonic acid, sulfonmethane,sulforidazine, sulfoxone, sulindac, sulisatin, sulisobenzone, sulmarin,sulmazole, suloctidil, sulphan blue, sulpiride, sultamicillin,sulthiame, sultopride, sultosilic acid, sumanirole, sumatriptan,SUN-N8075, suplatast, suprofen, suramin, surfactant TA, suriclone,suxibuzone, SYM-1010, SYM-2081, SYM-2207, symclosene, Syn-1253,Syn-2190, Syn-2869, synephrine, syrosingopine, T-1095, T-1249, T-3912,T-588, T-67, T-82, TA-2005, TA-2005, TA-993, tabimorelin, tacalcitol,tacedinaline, tacrine, tacrolimus, tadalafil, tafenoquine, tafluposide,TAK-375, TAK-427, TAK-559, taka-diastase, talampanel, talampicillin,talaporfin, talastine, talbutal, talinolol, talipexole, talnetant,talniflumate, taltirelin, tamoxifen, tamsulosin, tandospirone,tannoform, taprostene, tariquidar, TAS-103, tasosartan, taurocholicacid, taurolidine, tazanolast, tazarotene, tazobactam,tazobactam+piperacillin, TBC-3711, TCH-346, tebipenem, teboroxime,tecadenoson, tecastemizole, Technetium ⁹⁹Tc, teclothiazide, teclozan,tedisamil, teflurane, tegafur, tegafur+uracil, tegaserod, teicoplanin,telbivudine, telenzepine, telithromycin, telmesteine, telmisartan,telomerase inhibs, temazepam, temiverine, temocapril, temocillin,temoporfin, temozolomide, tenatoprazole, tenecteplase, tenidap,teniposide, tenofovir, tenofovir disoproxil, tenonitrozole, tenoxicam,tenuazonic acid, teprenone, terazosin, terbinafine, terbutaline,terconazole, terfenadine, terguride, terlipressin, terodiline,terofenamate, terpin, tertalolol, tert-pentyl alcohol, tesaglitazar,tesmilifene, testolactone, testosterone, tetrabamate, tetrabarbital,tetrabenazine, tetracaine, tetrachloroethylene, tetracine, tetracycline,tetrahydrozoline, tetrandrine, tetrantoin, tetrazepam, tetrofosmin,tetroxoprim, Tevenel®, tezacitabine, tezosentan, thalidomide,thenaldine, thenyldiamine, theobromine, theofibrate, theophylline,thiabendazole, thiacetazone, thiacymserine, thialbarbital, thiamine,thiamiprine, thiamphenicol, thiamylal, thiazesim, thiazinamium,thiazolinobutazone, thiazolsulfone, thibenzazoline, thiemalat,thiethylperazine, thimerfonate, thimerosal, thiobarbital,thiobutabarbital, thiocarbamizine, thiocarbarsone, thiocolchicine,thiocresol, thioctic acid, thioglycerol, thioguanine, thioimrag,thiopental, thiophosphoramide, thiopropazate, thioproperazine,thioridazine, thiosulfate, thiothixene, thiovir, thiphenamil, thiram,thonzylamine, thozalinone, thromboplastin, thurfyl nicotinate,thymectacin, thymol, thymopentin, thymyl N-isoamylcarbamate, thyropropicacid, thyroxine, tiadenol, tiagabine, tiamenidine, tianeptine, tiapride,tiaprofenic acid, tiaramide, tiazofurin, tibezonium, tibolone,ticarcillin, ticlopidine, ticrynafen, tiemonium, tigecycline, tigemonam,tigloidine, tilidine, tilisolol, tilmacoxib, tiludronic acid, timentin,timepidium, timiperone, timolol, timonacic, tin ethyl etiopurpurin,tinazoline, tinidazole, tinoridine, tiocarlide, tioclomarol,tioconazole, tiopronin, tiotropium, tioxolone, tipepidine, tipifarnib,tipranavir, tiquizium, tirapazamine, tiratricol, tirilazad, tirofiban,tiropramide, titanium sulfate, tiuxetan, tixocortol, tizanidine,TLK-199, TLK-286, TNF-β analogue, TNP-470, TO-186, tobramycin,tocainide, tocamphyl, tocladesine, tocoretinate, todralazine, tofenacin,tofimilast, tofisopam, tolazamid, tolazolin, tolbutamide, tolcapone,tolciclate, tolcyclamide, tolevamer, tolfenamic acid, tolindate,toliprolol, tolmetin, tolnaftate, tolonidine, tolonium, toloxatone,tolperisone, tolpropamine, tolrestat, tolserine, tolterodine, tolvaptan,tolycaine, topiramate, topoisomerase, topotecan, torasemide, torcetapib,torcitabine, toremifene, torsemide, tositumomab, tosulfloxacin,tramadol, tramazoline, trandolapril, tranexamic acid, tranilast,trans-retinoic acid, tranylcypromine, trapidil, trastuzumab, travoprost,traxanox, traxoprodil, trazodone, tremacamra, trenbolone, trengestone,treosulfan, trepibutone, treprostinol, tretinoin, tretoquinol, TRH,TRI-50b, triacetin, triamcinolone, triamcinolone, triamcinolone,triamcinolone acetonide, triamterene, triapine, triaziquone, triazolam,tribenoside, tribromophenate, trichlorfon, trichlormethiazide,trichlormethine, trichloroethylene, triclobisonium, triclocarban,triclofenol piperazine, triclofos, triclosan, tricromyl, tridihexethyliodide, trientine, triethanolamine, triethylenemelamine,trifluoperazine, trifluperidol, triflupromazine, trifluridine,triflusal, triflutate, trihexyphenidyl, trimazosin, trimebutine,trimecaine, trimeprazine, trimetazidine, trimethadione, trimethaphan,trimethobenzamide, trimethoprim, trimetozine, trimetrexate,trimipramine, trimoprostil, triolstane, trioxsalen, tripamide,triparanol, tripelennamine, triprolidine, triptorelin, tritiozine,tritoqualine, TRK-530, TRK-820, troclosene, trofosfamide, troglitazone,troleandomycin, trolnitrate, tromantadine, trometamol, trometamol,tromethamine, tromethamine, tropacine, tropesin, tropicamide, tropine,tropisetron, trospectomycin, trospium, trovafloxacin, troxacitabine,troxerutin, troxipide, trypan red, tryparsamide, tryptophan, TSH,TSN-09, TU-2100, tuaminoheptane, tubercidin, tubocurarine chloride,tulobuterol, TV-3326, TY-11223, TY-12533, TYB-3215, tybamate, tyloxapol,tymazoline, tyramine, tyropanoate, ubenimex, ufenamate, undecylenicacid, unoprostone, UR-8880, uracil mustard, uralyt-U, urapidil, urea,uredepa, urethan, uridine 5′-triphosphate, urinastatin, ursodeoxycholicacid, ursodiol, ushercell, uzarin, vaccine, Diphtheria Vaccine,Polyvalent Vaccine, valacyclovir, valdecoxib, valdetamide, valethamate,valganciclovir, valnoctamide, valomaciclovir, valproate, valproic acid,valpromide, valrocemide, valrubicin, valsartan, valspodar, vardenafil,varespladib, varicella virus, vatanidipine, VEA, vecuronium, velnacrine,venlafaxine, veralipride, verapamil, verteporfin, vesnarinone,vetrabutine, VF-233, VI-0134, vidarabine, vigabatrin, vilazodone,viloxazine, viminol, vinbarbital, vinblastine, vinburnine, vincamine,vinconate, vincristine, vindesine, vinflunine, vinorelbine, vinpocetine,vinyl ether, vinylbital, viquidil, viridin, visnadine, vitamin A,vitamin B12, vitamin C, vitamin D2, vitamin D3, vitamin K5, prenatalvitamins, VLA-4 antagonists, VNP-4010M, voglibose, voriconazole,vorozole, VUF-K-8788, warfarin, WF-10, WMC-79, wound healing matrix,WP-170, xaliproden, xamoterol, xanomeline, xanthinol niacinate,xemilofiban, xenbucin, xibenolol, xibornol, ximelagatran, ximoprofen,xipamide, xorphanol, XR-5118, XR-5944, xylometazoline, xylose, YH-1885,YM-511, YM-598, yohimbine, YT-146, Z-321, Z-335, zafirlukast,zalcitabine, zaldaride, zaleplon, zaltoprofen, zanamivir, zanapezil,zatebradine, ZD-0473, ZD-0947, ZD-6126, ZD-9331, zebularine, zelandopam,zenarestat, ziconotide, zidovudine, zileuton, zimeldine, zinc acetate,zinc acexamate, zinc ibuprofenate, zinc p-phenolsulfonate, zincsalicylate, zinostatin, zinostatin stimalamer, zipeprol, ziprasidone,zofenopril, zofenpril+HCTZ, zoledronic acid, zolimidine, zolmitriptan,zolpidem, zomepirac, zonampanel, zoniporide, zonisamide, zopiclone,zopolrestat, zorubicin, zosuquidar, zotepine, ZP-123, Z-tamoxifen,zuclopenthixol, α1-antitrypsin, α-bisabolol, α-chloralose, α-ethylbenzylalcohol, α-glucose-1-phosphate, α-phenylbutyramide, α-santonin,α-terpineol, α-tocopherol, β-alethine, β-benzalbutyramide, β-carotene,β-eucaine, β-propiolactone, β-sitosterol, γ-aminobutyric acid,γ-hydroxybutyrate, γ-linolenic acid, δ-aminolevulinic acid,ε-acetamidocaproic, and ε-aminocaproic acid. See also U.S. Pat. No.7,927,613, which is incorporated herein by reference in its entirety.Other pharmaceutically acceptable coformers include those delineated inthe “Generally Regarded as Safe” (“GRAS”) and/or the US FDA “EverythingAdded to Food in the United States” (“EAFUS”) lists.

In some embodiments, at least one of the one or more pharmaceuticallyacceptable coformers is niclosamide or a pharmaceutically acceptablesalt or hydrate thereof; or a niclosamide analog, or a pharmaceuticallyacceptable salt or hydrate thereof. In some of these embodiments, atleast one of the one or more pharmaceutically acceptable coformers canbe a compound having any one of formulas (I), (XVIII)-(XXV), and XXVII,e.g., formula XXIV or XXV; or any one of the compounds delineated above.In certain of these embodiments, at least one of the one or morepharmaceutically acceptable coformers can be a niclosamide analoguehaving any one of formulas (I), (XVIII)-(XXV), and XXVII, e.g., formulaXXIV or XXV; or XXVI; or any one of the compounds specificallydelineated above. In certain of these embodiments, the chemical entitycan be a niclosamide or a pharmaceutically acceptable salt or hydratethereof (e.g., niclosamide).

Non-Limiting Combinations

In some embodiments, the cocrystal includes (i) niclosamide or aniclosamide analog; and (ii) a pharmaceutically acceptable salt and/orhydrate of niclosamide; or a pharmaceutically acceptable salt and/orhydrate of a niclosamide analog.

In some embodiments, the cocrystal includes (i) niclosamide; and (ii) apharmaceutically acceptable salt and/or hydrate of niclosamide; or apharmaceutically acceptable salt and/or hydrate of niclosamide of aniclosamide analog.

In some embodiments, the cocrystal includes (i) niclosamide or aniclosamide analog; and (ii) a second API.

In some embodiments, the cocrystal includes (i) a pharmaceuticallyacceptable salt and/or hydrate of niclosamide; or a pharmaceuticallyacceptable salt and/or hydrate of niclosamide of a niclosamide analog;and (ii) a second API.

In some embodiments, the cocrystal includes (i) niclosamide; and (ii) asecond API.

In some embodiments, the cocrystal includes (i) a pharmaceuticallyacceptable salt and/or hydrate of niclosamide; or a pharmaceuticallyacceptable salt and/or hydrate of niclosamide of a niclosamide analog;and (ii) an amino acid (e.g., proline, e.g., D-proline, or L-proline, orracemic proline).

In some embodiments, the cocrystal includes (i) niclosamide; and (ii) anamino acid (e.g., proline, e.g., D-proline, or L-proline, or racemicproline).

In some embodiments, the cocrystal includes (i) a pharmaceuticallyacceptable salt and/or hydrate of niclosamide; or a pharmaceuticallyacceptable salt and/or hydrate of niclosamide of a niclosamide analog;and (ii) a 5-10 (e.g., 5-9, 5-6, or 5) membered heteroaryl, e.g., anitrogen-containing heteroaryl, e.g., imidazole.

In some embodiments, the cocrystal includes (i) niclosamide; and (ii) a5-10 (e.g., 5-9, 5-6, or 5) membered heteroaryl, e.g., anitrogen-containing heteroaryl, e.g., imidazole.

For examples, see Sanphui, P. Cryst. Growth Des. 2012, 12, 4588;Imramovský, A. Crystals 2012, 2, 349-361; and Grifasi, F. Cryst. GrowthDes. 2015, 15, 4588.

Properties

In some embodiments, the resulting co-crystals confer enhanced and/ornew and beneficial properties to the chemical entity (and/or to one ormore of the conformers, e.g., when a conformer is a second API) ascompared to the chemical entity in a free form (including free acids,free bases, and zwitter ions, hydrates, solvates, etc.), or an acid orbase salt thereof particularly with respect to, e.g., solubility,dissolution, bioavailability, stability, Cmax, Tmax, permeabilityprocessability, therapeutic plasma concentration, hygroscopicity,localized concentration, crystallization of amorphous compounds,decrease in form diversity (including polymorphism and crystal habit),change in morphology or crystal habit.

In some embodiments, the cocrystals have an oral bioavailability (F) ofless than about 50%, or less than about 40%, or less than about 30%, orless than about 20%, or less than about 10%, or less than about 5%, orless than about 2%, or less than about 1%. In certain embodiments, thechemical entities described herein have an oral bioavailability (F) ofless than about 20%, e.g., less than about 19%, less than about 18%,less than about 17%, less than about 16%, less than about 15%, less thanabout 14%, less than about 13%, less than about 12%, less than about11%, less than about 10%, less than about 9%, less than about 8%, lessthan about 7%, less than about 6%, less than about 5%, less than about4%, less than about 3%, less than about 2%, less than about 1%, or lessthan about 0.5%.

In some embodiments, the cocrystals have a relatively low aqueoussolubility. Low aqueous solubility refers to a compound having asolubility in water which is less than or equal to 10 mg/mL, whenmeasured at 20° C. In certain embodiments, the chemical entitiesdescribed herein have aqueous solubility of less than or equal to 900,800, 700, 600, 500, 400, 300, 200 150 100, 90, 80, 70, 60, 50, 40, 30,20 micrograms/mL, or further 10, 5 or 1 micrograms/mL, or further 900,800, 700, 600, 500, 400, 300, 200 150, 100 90, 80, 70, 60, 50, 40, 30,20, or 10 ng/mL, or less than 10 ng/mL when measured at 20° C.

In some embodiments, the cocrystals have a relatively low drugpermeability.

Pharmaceutical Compositions and Administration

General

A chemical entity (e.g., a compound exhibiting activity as amitochondrial uncoupling agent or a pharmaceutically acceptable saltand/or hydrate and/or cocrystal thereof; e.g., a compound, such asniclosamide or a pharmaceutically acceptable salt and/or hydrate and/orcocrystal thereof; e.g., a compound, such as a niclosamide analog, or apharmaceutically acceptable salt and/or hydrate and/or cocrystalthereof) is administered to a subject in need thereof by any route whichmakes the compound bioavailable (e.g., locally bioavailable).

In some embodiments, a chemical entity (e.g., a compound exhibitingactivity as a mitochondrial uncoupling agent or a pharmaceuticallyacceptable salt and/or hydrate and/or cocrystal thereof; e.g., acompound, such as niclosamide or a pharmaceutically acceptable saltand/or hydrate and/or cocrystal thereof; e.g., a compound, such as aniclosamide analog, or a pharmaceutically acceptable salt and/or hydrateand/or cocrystal thereof) is administered as a pharmaceuticalcomposition that includes the chemical entity and one or morepharmaceutically acceptable excipients, and optionally one or more othertherapeutic agents as described herein.

In some embodiments, the chemical entities can be administered incombination with one or more conventional pharmaceutical excipients.Pharmaceutically acceptable excipients include, but are not limited to,ion exchangers, alumina, aluminum stearate, lecithin, self-emulsifyingdrug delivery systems (SEDDS) such as d-α-tocopherol polyethylene glycol1000 succinate, surfactants used in pharmaceutical dosage forms such asTweens, poloxamers or other similar polymeric delivery matrices, serumproteins, such as human serum albumin, buffer substances such asphosphates, tris, glycine, sorbic acid, potassium sorbate, partialglyceride mixtures of saturated vegetable fatty acids, water, salts orelectrolytes, such as protamine sulfate, disodium hydrogen phosphate,potassium hydrogen phosphate, sodium-chloride, zinc salts, colloidalsilica, magnesium trisilicate, polyvinyl pyrrolidone, cellulose-basedsubstances, polyethylene glycol, sodium carboxymethyl cellulose,polyacrylates, waxes, polyethylene-polyoxypropylene-block polymers, andwool fat. Cyclodextrins such as α-, β, and γ-cyclodextrin, or chemicallymodified derivatives such as hydroxyalkylcyclodextrins, including 2- and3-hydroxypropyl-β-cyclodextrins, or other solubilized derivatives canalso be used to enhance delivery of compounds described herein. Dosageforms or compositions containing a chemical entity as described hereinin the range of 0.005% to 100% with the balance made up from non-toxicexcipient may be prepared. The contemplated compositions may contain0.001%-100% of a chemical entity provided herein, in one embodiment0.1-95%, in another embodiment 75-85%, in a further embodiment 20-80%.Actual methods of preparing such dosage forms are known, or will beapparent, to those skilled in this art; for example, see Remington: TheScience and Practice of Pharmacy, 22^(nd) Edition (Pharmaceutical Press,London, UK. 2012).

In some embodiments, the chemical entities described herein or apharmaceutical composition thereof can be administered to subject inneed thereof by any accepted route of administration. Acceptable routesof administration include, but are not limited to, buccal, cutaneous,endocervical, endosinusial, endotracheal, enteral, epidural,interstitial, intra-abdominal, intra-arterial, intrabronchial,intrabursal, intracerebral, intracisternal, intracoronary, intradermal,intraductal, intraduodenal, intradural, intraepidermal, intraesophageal,intragastric, intragingival, intraileal, intralymphatic, intramedullary,intrameningeal, intramuscular, intraovarian, intraperitoneal,intraprostatic, intrapulmonary, intrasinal, intraspinal, intrasynovial,intratesticular, intrathecal, intratubular, intratumor, intrauterine,intravascular, intravenous, nasal, nasogastric, oral, parenteral,percutaneous, peridural, rectal, respiratory (inhalation), subcutaneous,sublingual, submucosal, topical, transdermal, transmucosal,transtracheal, ureteral, urethral and vaginal.

Local Administration

In some embodiments, the chemical entities described herein or apharmaceutical composition thereof are suitable for localadministration, e.g., local administration by way of topicallyadministering the chemical entity or composition thereof at a particulartreatment site, (e.g., the digestive tract, the gastrointestinal (“GI”)tract, eye, joint, or skin) so as to provide local administration of thechemical entity to the area in need of treatment (e.g., oral cavity; GItract, e.g., the colon; eye; skin; or joint). In certain embodiments,minimal systemic exposure of the chemical entity occurs during saidlocal administration. Examples of such compositions include, withoutlimitation, compositions for rectal administration, oral administration,dermal administration, or implant. In certain embodiments, compositionsare for other than oral administration.

In some embodiments, the chemical entities described herein or apharmaceutical composition thereof are suitable for local administrationto the GI tract. In certain embodiments, upon administration, the localconcentration of the chemical entity in the GI tract is higher (e.g.,from about 2 times higher to about 50 times higher, from about 5 timeshigher to about 50 times higher; from about 5 times higher to about 25times higher; from about 5 times higher to about 15 times higher; e.g.,about 50 times higher, about 25 time higher, about 20 times higher,about 15 times higher, about 10 times higher, about 5 times higher,e.g., at least about 10 times higher) than the concentration of thechemical entity in the plasma compartment. In certain of theseembodiments, the chemical entity in the plasma compartment is subject tofirst pass metabolism.

In some embodiments, the chemical entities described herein or apharmaceutical composition thereof are suitable for local administrationto one or more specific locations within the digestive or GI tract. Forexample, at least some of the chemical entity is present in the upper GItract (e.g., stomach); or at least some of the agent is present in thelower GI tract (e.g., the large intestine, e.g., the colon, e.g., theascending colon and/or transverse colon and/or distal colon; or thesmall bowel). As a further example, at least some of the chemical entityis present in the ascending colon and/or the transverse colon and/or thedistal colon and/or the small bowel and/or the stomach. Methods of saidlocal administration can include, without limitation, rectaladministration and/or oral administration.

In certain embodiments, the chemical entities described herein or apharmaceutical composition thereof are suitable for local, topicaladministration to the digestive or GI tract, e.g., rectaladministration. Rectal compositions include, without limitation, enemas,rectal gels, rectal foams, rectal aerosols, suppositories, jellysuppositories, and enemas (e.g., retention enemas).

Pharmacologically acceptable excipients usable in the rectal compositionas a gel, cream, enema, or rectal suppository, include, withoutlimitation, any one or more of cocoa butter glycerides, syntheticpolymers such as polyvinylpyrrolidone, PEG (like PEG ointments),glycerine, glycerinated gelatin, hydrogenated vegetable oils,poloxamers, mixtures of polyethylene glycols of various molecularweights and fatty acid esters of polyethylene glycol Vaseline, anhydrouslanolin, shark liver oil, sodium saccharinate, menthol, sweet almondoil, sorbitol, sodium benzoate, anoxid SBN, vanilla essential oil,aerosol, parabens in phenoxyethanol, sodium methyl p-oxybenzoate, sodiumpropyl p-oxybenzoate, diethylamine, carbomers, carbopol,methyloxybenzoate, macrogol cetostearyl ether, cocoyl caprylocaprate,isopropyl alcohol, propylene glycol, liquid paraffin, xanthan gum,carboxy-metabisulfite, sodium edetate, sodium benzoate, potassiummetabisulfite, grapefruit seed extract, methyl sulfonyl methane (MSM),lactic acid, glycine, vitamins, such as vitamin A and E and potassiumacetate.

In certain embodiments, suppositories can be prepared by mixing thechemical entities described herein with suitable non-irritatingexcipients or carriers such as cocoa butter, polyethylene glycol or asuppository wax which are solid at ambient temperature but liquid atbody temperature and therefore melt in the rectum and release the activecompound. In other embodiments, compositions for rectal administrationare in the form of an enema.

Enema Formulations

In some embodiments, enema formulations containing the chemical entitiesdescribed herein are provided in “ready-to-use” form.

In some embodiments, enema formulations containing the chemical entitiesdescribed herein are provided in one or more kits or packs. In certainembodiments, the kit or pack includes two or more separatelycontained/packaged components, e.g. two components, which when mixedtogether, provide the desired formulation (e.g., as a suspension). Incertain of these embodiments, the two component system includes a firstcomponent and a second component, in which: (i) the first component(e.g., contained in a sachet) includes the chemical entity (as describedanywhere herein) and optionally one or more pharmaceutically acceptableexcipients (e.g., together formulated as a solid preparation, e.g.,together formulated as a wet granulated solid preparation); and (ii) thesecond component (e.g., contained in a vial or bottle) includes one ormore liquids and optionally one or more other pharmaceuticallyacceptable excipients together forming a liquid carrier. Prior to use(e.g., immediately prior to use), the contents of (i) and (ii) arecombined to form the desired enema formulation, e.g., as a suspension.In other embodiments, each of component (i) and (ii) is provided in itsown separate kit or pack.

In some embodiments, each of the one or more liquids is water, or aphysiologically acceptable solvent, or a mixture of water and one ormore physiologically acceptable solvents. Typical such solvents include,without limitation, glycerol, ethylene glycol, propylene glycol,polyethylene glycol and polypropylene glycol. In certain embodiments,each of the one or more liquids is water. In other embodiments, each ofthe one or more liquids is an oil, e.g. natural and/or synthetic oilsthat are commonly used in pharmaceutical preparations.

Further pharmaceutical excipients and carriers that may be used in thepharmaceutical products herein described are listed in various handbooks(e.g. D. E. Bugay and W. P. Findlay (Eds) Pharmaceutical excipients(Marcel Dekker, New York, 1999), E-M Hoepfner, A. Reng and P. C. Schmidt(Eds) Fiedler Encyclopedia of Excipients for Pharmaceuticals, Cosmeticsand Related Areas (Edition Cantor, Munich, 2002) and H. P. Fielder (Ed)Lexikon der Hilfsstoffe fiir Pharmazie, Kosmetik and angrenzende Gebiete(Edition Cantor Aulendorf, 1989)).

In some embodiments, each of the one or more pharmaceutically acceptableexcipients can be independently selected from thickeners, viscosityenhancing agents, bulking agents, mucoadhesive agents, penetrationenhancers, buffers, preservatives, diluents, binders, lubricants,glidants, disintegrants, fillers, solubilizing agents, pH modifyingagents, preservatives, stabilizing agents, anti-oxidants, wetting oremulsifying agents, suspending agents, pigments, colorants, isotonicagents, chelating agents, emulsifiers, and diagnostic agents.

In certain embodiments, each of the one or more pharmaceuticallyacceptable excipients can be independently selected from thickeners,viscosity enhancing agents, mucoadhesive agents, buffers, preservatives,diluents, binders, lubricants, glidants, disintegrants, and fillers.

In certain embodiments, each of the one or more pharmaceuticallyacceptable excipients can be independently selected from thickeners,viscosity enhancing agents, bulking agents, mucoadhesive agents,buffers, preservatives, and fillers.

In certain embodiments, each of the one or more pharmaceuticallyacceptable excipients can be independently selected from diluents,binders, lubricants, glidants, and disintegrants.

Examples of thickeners, viscosity enhancing agents, and mucoadhesiveagents include without limitation: gums, e.g. xanthan gum, guar gum,locust bean gum, tragacanth gums, karaya gum, ghatti gum, cholla gum,psyllium seed gum and gum arabic; poly(carboxylic acid-containing) basedpolymers, such as poly (acrylic, maleic, itaconic, citraconic,hydroxyethyl methacrylic or methacrylic) acid which have stronghydrogen-bonding groups, or derivatives thereof such as salts andesters; cellulose derivatives, such as methyl cellulose, ethylcellulose, methylethyl cellulose, hydroxymethyl cellulose, hydroxyethylcellulose, hydroxypropyl cellulose, hydroxyethyl ethyl cellulose,carboxymethyl cellulose, hydroxypropylmethyl cellulose or celluloseesters or ethers or derivatives or salts thereof; clays such asmanomorillonite clays, e.g. Veegun, attapulgite clay; polysaccharidessuch as dextran, pectin, amylopectin, agar, mannan or polygalactonicacid or starches such as hydroxypropyl starch or carboxymethyl starch;polypeptides such as casein, gluten, gelatin, fibrin glue; chitosan,e.g. lactate or glutamate or carboxymethyl chitin; glycosaminoglycanssuch as hyaluronic acid; metals or water soluble salts of alginic acidsuch as sodium alginate or magnesium alginate; schleroglucan; adhesivescontaining bismuth oxide or aluminium oxide; atherocollagen; polyvinylpolymers such as carboxyvinyl polymers; polyvinylpyrrolidone (povidone);polyvinyl alcohol; polyvinyl acetates, polyvinylmethyl ethers, polyvinylchlorides, polyvinylidenes, and/or the like; polycarboxylated vinylpolymers such as polyacrylic acid as mentioned above; polysiloxanes;polyethers; polyethylene oxides and glycols; polyalkoxys andpolyacrylamides and derivatives and salts thereof. Preferred examplescan include cellulose derivatives, such as methyl cellulose, ethylcellulose, methylethyl cellulose, hydroxymethyl cellulose, hydroxyethylcellulose, hydroxypropyl cellulose, hydroxyethyl ethyl cellulose,carboxymethyl cellulose, hydroxypropylmethyl cellulose or celluloseesters or ethers or derivatives or salts thereof (e.g., methylcellulose); and polyvinyl polymers such as polyvinylpyrrolidone(povidone).

Examples of preservatives include without limitation: benzalkoniumchloride, benzoxonium chloride, benzethonium chloride, cetrimide,sepazonium chloride, cetylpyridinium chloride, domiphen bromide(Bradosol®), thiomersal, phenylmercuric nitrate, phenylmercuric acetate,phenylmercuric borate, methylparaben, propylparaben, chlorobutanol,benzyl alcohol, phenyl ethyl alcohol, chlorohexidine, polyhexamethylenebiguanide, sodium perborate, imidazolidinyl urea, sorbic acid, Purite®),Polyquart®), and sodium perborate tetrahydrate and the like.

In certain embodiments, the preservative is a paraben, or apharmaceutically acceptable salt thereof. In some embodiments, theparaben is an alkyl substituted 4-hydroxybenzoate, or a pharmaceuticallyacceptable salt or ester thereof. In certain embodiments, the alkyl is aC1-C4 alkyl. In certain embodiments, the preservative is methyl4-hydroxybenzoate (methylparaben), or a pharmaceutically acceptable saltor ester thereof, propyl 4-hydroxybenzoate (propylparaben), or apharmaceutically acceptable salt or ester thereof, or a combinationthereof.

Examples of buffers include without limitation: phosphate buffer system(sodium dihydrogen phosphate dehydrate, disodium phosphatedodecahydrate, bibasic sodium phosphate, anhydrous monobasic sodiumphosphate), bicarbonate buffer system, and bisulfate buffer system.

Examples of disintegrants include, without limitation: carmellosecalcium, low substituted hydroxypropyl cellulose (L-HPC), carmellose,croscarmellose sodium, partially pregelatinized starch, dry starch,carboxymethyl starch sodium, crospovidone, polysorbate 80(polyoxyethylenesorbitan oleate), starch, sodium starch glycolate,hydroxypropyl cellulose pregelatinized starch, clays, cellulose,alginine, gums or cross linked polymers, such as cross-linked PVP(Polyplasdone XL from GAF Chemical Corp). In certain embodiments, thedisintegrant is crospovidone.

Examples of glidants and lubricants (aggregation inhibitors) includewithout limitation: talc, magnesium stearate, calcium stearate,colloidal silica, stearic acid, aqueous silicon dioxide, syntheticmagnesium silicate, fine granulated silicon oxide, starch, sodiumlaurylsulfate, boric acid, magnesium oxide, waxes, hydrogenated oil,polyethylene glycol, sodium benzoate, stearic acid glycerol behenate,polyethylene glycol, and mineral oil. In certain embodiments, theglidant/lubricant is magnesium stearate, talc, and/or colloidal silica;e.g., magnesium stearate and/or talc.

Examples of diluents, also referred to as “fillers” or “bulking agents”include without limitation: dicalcium phosphate dihydrate, calciumsulfate, lactose (e.g., lactose monohydrate), sucrose, mannitol,sorbitol, cellulose, microcrystalline cellulose, kaolin, sodiumchloride, dry starch, hydrolyzed starches, pregelatinized starch,silicone dioxide, titanium oxide, magnesium aluminum silicate andpowdered sugar. In certain embodiments, the diluent is lactose (e.g.,lactose monohydrate).

Examples of binders include without limitation: starch, pregelatinizedstarch, gelatin, sugars (including sucrose, glucose, dextrose, lactoseand sorbitol), polyethylene glycol, waxes, natural and synthetic gumssuch as acacia tragacanth, sodium alginate cellulose, includinghydroxypropylmethylcellulose, hydroxypropylcellulose, ethylcellulose,and veegum, and synthetic polymers such as acrylic acid and methacrylicacid copolymers, methacrylic acid copolymers, methyl methacrylatecopolymers, aminoalkyl methacrylate copolymers, polyacrylicacid/polymethacrylic acid and polyvinylpyrrolidone (povidone). Incertain embodiments, the binder is polyvinylpyrrolidone (povidone).

In some embodiments, enema formulations containing the chemical entitiesdescribed herein include water and one or more (e.g., all) of thefollowing excipients:

-   -   One or more (e.g., one, two, or three) thickeners, viscosity        enhancing agents, binders, and/or mucoadhesive agents (e.g.,        cellulose or cellulose esters or ethers or derivatives or salts        thereof (e.g., methyl cellulose); and polyvinyl polymers such as        polyvinylpyrrolidone (povidone);    -   One or more (e.g., one or two; e.g., two) preservatives, such as        a paraben, e.g., methyl 4-hydroxybenzoate (methylparaben), or a        pharmaceutically acceptable salt or ester thereof, propyl        4-hydroxybenzoate (propylparaben), or a pharmaceutically        acceptable salt or ester thereof, or a combination thereof;    -   One or more (e.g., one or two; e.g., two) buffers, such as        phosphate buffer system (e.g., sodium dihydrogen phosphate        dehydrate, disodium phosphate dodecahydrate);    -   One or more (e.g., one or two, e.g., two) glidants and/or        lubricants, such as magnesium stearate and/or talc;    -   One or more (e.g., one or two; e.g., one) disintegrants, such as        crospovidone; and    -   One or more (e.g., one or two; e.g., one) diluents, such as        lactose (e.g., lactose monohydrate).

In certain of these embodiments, the chemical entity is niclosamide, ora pharmaceutically acceptable salt and/or hydrate and/or cocrystalthereof; e.g., niclosamide.

In certain embodiments, enema formulations containing the chemicalentities described herein include water, methyl cellulose, povidone,methylparaben, propylparaben, sodium dihydrogen phosphate dehydrate,disodium phosphate dodecahydrate, crospovidone, lactose monohydrate,magnesium stearate, and talc. In certain of these embodiments, thechemical entity is niclosamide, or a pharmaceutically acceptable saltand/or hydrate and/or cocrystal thereof; e.g., niclosamide.

In certain embodiments, enema formulations containing the chemicalentities described herein are provided in one or more kits or packs. Incertain embodiments, the kit or pack includes two separatelycontained/packaged components, which when mixed together, provide thedesired formulation (e.g., as a suspension). In certain of theseembodiments, the two component system includes a first component and asecond component, in which: (i) the first component (e.g., contained ina sachet) includes the chemical entity (as described anywhere herein)and one or more pharmaceutically acceptable excipients (e.g., togetherformulated as a solid preparation, e.g., together formulated as a wetgranulated solid preparation); and (ii) the second component (e.g.,contained in a vial or bottle) includes one or more liquids and one ormore one or more other pharmaceutically acceptable excipients togetherforming a liquid carrier. In other embodiments, each of component (i)and (ii) is provided in its own separate kit or pack.

In certain of these embodiments, component (i) includes the chemicalentity (e.g., niclosamide, or a pharmaceutically acceptable salt and/orhydrate and/or cocrystal thereof; e.g., niclosamide) and one or more(e.g., all) of the following excipients:

-   -   (a) One or more (e.g., one) binders (e.g., a polyvinyl polymer,        such as polyvinylpyrrolidone (povidone);    -   (b) One or more (e.g., one or two, e.g., two) glidants and/or        lubricants, such as magnesium stearate and/or talc;    -   (c) One or more (e.g., one or two; e.g., one) disintegrants,        such as crospovidone; and    -   (d) One or more (e.g., one or two; e.g., one) diluents, such as        lactose (e.g., lactose monohydrate).

In certain embodiments, component (i) includes from about 40 weightpercent to about 80 weight percent (e.g., from about 50 weight percentto about 70 weight percent, from about 55 weight percent to about 70weight percent; from about 60 weight percent to about 65 weight percent;e.g., about 62.1 weight percent) of the chemical entity (e.g.,niclosamide, or a pharmaceutically acceptable salt and/or hydrate and/orcocrystal thereof; e.g., niclosamide).

In certain embodiments, component (i) includes from about 0.5 weightpercent to about 5 weight percent (e.g., from about 1.5 weight percentto about 4.5 weight percent, from about 2 weight percent to about 3.5weight percent; e.g., about 2.76 weight percent) of the binder (e.g.,povidone).

In certain embodiments, component (i) includes from about 0.5 weightpercent to about 5 weight percent (e.g., from about 0.5 weight percentto about 3 weight percent, from about 1 weight percent to about 3 weightpercent; about 2 weight percent e.g., about 1.9 weight percent) of thedisintegrant (e.g., crospovidone).

In certain embodiments, component (i) includes from about 10 weightpercent to about 50 weight percent (e.g., from about 20 weight percentto about 40 weight percent, from about 25 weight percent to about 35weight percent; e.g., about 31.03 weight percent) of the diluent (e.g.,lactose, e.g., lactose monohydrate).

In certain embodiments, component (i) includes from about 0.05 weightpercent to about 5 weight percent (e.g., from about 0.05 weight percentto about 3 weight percent) of the glidants and/or lubricants.

In certain embodiments (e.g., when component (i) includes one or morelubricants, such as magnesium stearate), component (i) includes fromabout 0.05 weight percent to about 1 weight percent (e.g., from about0.05 weight percent to about 1 weight percent; from about 0.1 weightpercent to about 1 weight percent; from about 0.1 weight percent toabout 0.5 weight percent; e.g., about 0.27 weight percent) of thelubricant (e.g., magnesium stearate).

In certain embodiments (when component (i) includes one or morelubricants, such as talc), component (i) includes from about 0.5 weightpercent to about 5 weight percent (e.g., from about 0.5 weight percentto about 3 weight percent, from about 1 weight percent to about 3 weightpercent; from about 1.5 weight percent to about 2.5 weight percent; fromabout 1.8 weight percent to about 2.2 weight percent; about 1.93 weightpercent) of the lubricant (e.g., talc).

In certain of these embodiments, each of (a), (b), (c), and (d) above ispresent.

In certain embodiments, component (i) includes the ingredients andamounts as shown in Table 7

TABLE 7 Ingredient Weight Percent niclosamide 40 weight percent to about80 weight percent (e.g., from about 50 weight percent to about 70 weightpercent, from about 55 weight percent to about 70 weight percent; fromabout 60 weight percent to about 65 weight percent; e.g., about 62.1weight percent) Crospovidone (Kollidon CL) 0.5 weight percent to about 5weight percent (e.g., from about 0.5 weight percent to about 3 weightpercent, from about 1 weight percent to about 3 weight percent; about1.93 weight percent lactose monohydrate about 10 weight percent to about50 weight (Pharmatose 200M) percent (e.g., from about 20 weight percentto about 40 weight percent, from about 25 weight percent to about 35weight percent; e.g., about 31.03 weight percent Povidone (Kollidon K30)about 0.5 weight percent to about 5 weight percent (e.g., from about 1.5weight percent to about 4.5 weight percent, from about 2 weight percentto about 3.5 weight percent; e.g., about 2.76 weight percent talc 0.5weight percent to about 5 weight percent (e.g., from about 0.5 weightpercent to about 3 weight percent, from about 1 weight percent to about3 weight percent; from about 1.5 weight percent to about 2.5 weightpercent; from about 1.8 weight percent to about 2.2 weight percent;e.g., about 1.93 weight percent Magnesium stearate about 0.05 weightpercent to about 1 weight percent (e.g., from about 0.05 weight percentto about 1 weight percent; from about 0.1 weight percent to about 1weight percent; from about 0.1 weight percent to about 0.5 weightpercent; e.g., about 0.27 weight percent

In certain embodiments, component (i) includes the ingredients andamounts as shown in Table 8.

TABLE 8 Ingredient Weight Percent niclosamide About 62.1 weight percent)Crospovidone (Kollidon CL) About 1.93 weight percent lactose monohydrateAbout 31.03 weight percent (Pharmatose 200M) Povidone (Kollidon K30)About 2.76 weight percent talc About 1.93 weight percent Magnesiumstearate About 0.27 weight percent

In certain embodiments, component (i) is formulated as a wet granulatedsolid preparation. In certain of these embodiments an internal phase ofingredients (the chemical entity, disintegrant, and diluent) arecombined and mixed in a high-shear granulator. A binder (e.g., povidone)is dissolved in water to form a granulating solution. This solution isadded to the Inner Phase mixture resulting in the development ofgranules. While not wishing to be bound by theory, granule developmentis believed to be facilitated by the interaction of the polymeric binderwith the materials of the internal phase. Once the granulation is formedand dried, an external phase (e.g., one or more lubricants—not anintrinsic component of the dried granulation), is added to the drygranulation. It is believed that lubrication of the granulation isimportant to the flowability of the granulation, in particular forpackaging. See, e.g., Example 8.

In certain of the foregoing embodiments, component (ii) includes waterand one or more (e.g., all) of the following excipients:

-   -   (a′) One or more (e.g., one, two; e.g., two) thickeners,        viscosity enhancing agents, binders, and/or mucoadhesive agents        (e.g., cellulose or cellulose esters or ethers or derivatives or        salts thereof (e.g., methyl cellulose); and polyvinyl polymers        such as polyvinylpyrrolidone (povidone);    -   (b′) One or more (e.g., one or two; e.g., two) preservatives,        such as a paraben, e.g., methyl 4-hydroxybenzoate        (methylparaben), or a pharmaceutically acceptable salt or ester        thereof, propyl 4-hydroxybenzoate (propylparaben), or a        pharmaceutically acceptable salt or ester thereof, or a        combination thereof; and    -   (c′) One or more (e.g., one or two; e.g., two) buffers, such as        phosphate buffer system (e.g., sodium dihydrogen phosphate        dihydrate, disodium phosphate dodecahydrate);

In certain of the foregoing embodiments, component (ii) includes waterand one or more (e.g., all) of the following excipients:

-   -   (a″) a first thickener, viscosity enhancing agent, binder,        and/or mucoadhesive agent (e.g., a cellulose or cellulose ester        or ether or derivative or salt thereof (e.g., methyl        cellulose));    -   (a′″) a second thickener, viscosity enhancing agent, binder,        and/or mucoadhesive agent (e.g., a polyvinyl polymer, such as        polyvinylpyrrolidone (povidone));    -   (b″) a first preservative, such as a paraben, e.g., propyl        4-hydroxybenzoate (propylparaben), or a pharmaceutically        acceptable salt or ester thereof;    -   (b″) a second preservative, such as a paraben, e.g., methyl        4-hydroxybenzoate (methylparaben), or a pharmaceutically        acceptable salt or ester thereof,    -   (c″) a first buffer, such as phosphate buffer system (e.g.,        disodium phosphate dodecahydrate);    -   (c′″) a second buffer, such as phosphate buffer system (e.g.,        sodium dihydrogen phosphate dehydrate),

In certain embodiments, component (ii) includes from about 0.05 weightpercent to about 5 weight percent (e.g., from about 0.05 weight percentto about 3 weight percent, from about 0.1 weight percent to about 3weight percent; e.g., about 1.4 weight percent) of (a″).

In certain embodiments, component (ii) includes from about 0.05 weightpercent to about 5 weight percent (e.g., from about 0.05 weight percentto about 3 weight percent, from about 0.1 weight percent to about 2weight percent; e.g., about 1.0 weight percent) of (a′″).

In certain embodiments, component (ii) includes from about 0.005 weightpercent to about 0.1 weight percent (e.g., from about 0.005 weightpercent to about 0.05 weight percent; e.g., about 0.02 weight percent)of (b″).

In certain embodiments, component (ii) includes from about 0.05 weightpercent to about 1 weight percent (e.g., from about 0.05 weight percentto about 0.5 weight percent; e.g., about 0.20 weight percent) of (b′″).

In certain embodiments, component (ii) includes from about 0.05 weightpercent to about 1 weight percent (e.g., from about 0.05 weight percentto about 0.5 weight percent; e.g., about 0.15 weight percent) of (c″).

In certain embodiments, component (ii) includes from about 0.005 weightpercent to about 0.5 weight percent (e.g., from about 0.005 weightpercent to about 0.3 weight percent; e.g., about 0.15 weight percent) of(c′″).

In certain of these embodiments, each of (a″)-(c′″) is present.

In certain embodiments, component (ii) includes water (up to 100%) andthe ingredients and amounts as shown in Table 9.

TABLE 9 Ingredient Weight Percent methyl cellulose (Methocel 0.05 weightpercent to about 5 weight A15C premium) percent (e.g., from about 0.05weight percent to about 3 weight percent, from about 0.1 weight percentto about 3 weight percent; e.g., about 1.4 weight percent Povidone(Kollidon K30) 0.05 weight percent to about 5 weight percent (e.g., fromabout 0.05 weight percent to about 3 weight percent, from about 0.1weight percent to about 2 weight percent; e.g., about 1.0 weight percentpropyl 4-hydroxybenzoate about 0.005 weight percent to about 0.1 weightpercent (e.g., from about 0.005 weight percent to about 0.05 weightpercent; e.g., about 0.02 weight percent) methyl 4-hydroxybenzoate about0.05 weight percent to about 1 weight percent (e.g., from about 0.05weight percent to about 0.5 weight percent; e.g., about 0.20 weightpercent) disodium phosphate about 0.05 weight percent to about 1dodecahydrate weight percent (e.g., from about 0.05 weight percent toabout 0.5 weight percent; e.g., about 0.15 weight percent) sodiumdihydrogen phospahate about 0.005 weight percent to about dihydrate 0.5weight percent (e.g., from about 0.005 weight percent to about 0.3weight percent; e.g., about 0.15 weight percent)

In certain embodiments, component (ii) includes water (up to 100%) andthe ingredients and amounts as shown in Table 10.

TABLE 10 Ingredient Weight Percent methyl cellulose (Methocel A15C about1.4 weight percent premium) Povidone (Kollidon K30) about 1.0 weightpercent propyl 4-hydroxybenzoate about 0.02 weight percent methyl4-hydroxybenzoate about 0.20 weight percent disodium phosphatedodecahydrate about 0.15 weight percent sodium dihydrogen phospahatedihydrate about 0.15 weight percent

Ready-to-use” enemas are generally be provided in a “single-use” sealeddisposable container of plastic or glass. Those formed of a polymericmaterial preferably have sufficient flexibility for ease of use by anunassisted patient. Typical plastic containers can be made ofpolyethylene. These containers may comprise a tip for directintroduction into the rectum. Such containers may also comprise a tubebetween the container and the tip. The tip is preferably provided with aprotective shield which is removed before use. Optionally the tip has alubricant to improve patient compliance.

In some embodiments, the enema formulation (e.g., suspension) is pouredinto a bottle for delivery after it has been prepared in a separatecontainer. In certain embodiments, the bottle is a plastic bottle (e.g.,flexible to allow for delivery by squeezing the bottle), which can be apolyethylene bottle (e.g., white in color). In some embodiments, thebottle is a single chamber bottle, which contains the suspension orsolution. In other embodiments, the bottle is a multichamber bottle,where each chamber contains a separate mixture or solution. In stillother embodiments, the bottle can further include a tip or rectalcannula for direct introduction into the rectum. In some embodiments,the enema formulation can be delivered in the device shown in FIGS.3A-3C, which includes a plastic bottle, a breakable capsule, and arectal cannula and single flow pack.

Oral Delivery

In other embodiments, the chemical entities described herein or apharmaceutical composition thereof are suitable for local delivery tothe digestive or GI tract by way of oral administration (e.g., solid orliquid dosage forms.).

Solid dosage forms for oral administration include capsules, tablets,pills, powders, and granules. In such solid dosage forms, the chemicalentity is mixed with one or more pharmaceutically acceptable excipients,such as sodium citrate or dicalcium phosphate and/or: a) fillers orextenders such as starches, lactose, sucrose, glucose, mannitol, andsilicic acid, b) binders such as, for example, carboxymethylcellulose,alginates, gelatin, polyvinylpyrrolidinone, sucrose, and acacia, c)humectants such as glycerol, d) disintegrating agents such as agar-agar,calcium carbonate, potato or tapioca starch, alginic acid, certainsilicates, and sodium carbonate, e) solution retarding agents such asparaffin, f) absorption accelerators such as quaternary ammoniumcompounds, g) wetting agents such as, for example, cetyl alcohol andglycerol monostearate, h) absorbents such as kaolin and bentonite clay,and i) lubricants such as talc, calcium stearate, magnesium stearate,solid polyethylene glycols, sodium lauryl sulfate, and mixtures thereof.In the case of capsules, tablets and pills, the dosage form may alsocomprise buffering agents. Solid compositions of a similar type may alsobe employed as fillers in soft and hard-filled gelatin capsules usingsuch excipients as lactose or milk sugar as well as high molecularweight polyethylene glycols and the like.

In one embodiment, the compositions will take the form of a unit dosageform such as a pill or tablet and thus the composition may contain,along with a chemical entity provided herein, a diluent such as lactose,sucrose, dicalcium phosphate, or the like; a lubricant such as magnesiumstearate or the like; and a binder such as starch, gum acacia,polyvinylpyrrolidine, gelatin, cellulose, cellulose derivatives or thelike. In another solid dosage form, a powder, marume, solution orsuspension (e.g., in propylene carbonate, vegetable oils, PEG's,poloxamer 124 or triglycerides) is encapsulated in a capsule (gelatin orcellulose base capsule). Unit dosage forms in which one or more chemicalentities provided herein or additional active agents are physicallyseparated are also contemplated; e.g., capsules with granules (ortablets in a capsule) of each drug; two-layer tablets; two-compartmentgel caps, etc. Enteric coated or delayed release oral dosage forms arealso contemplated.

Other physiologically acceptable compounds include wetting agents,emulsifying agents, dispersing agents or preservatives that areparticularly useful for preventing the growth or action ofmicroorganisms. Various preservatives are well known and include, forexample, phenol and ascorbic acid.

In certain embodiments the excipients are sterile and generally free ofundesirable matter. These compositions can be sterilized byconventional, well-known sterilization techniques. For various oraldosage form excipients such as tablets and capsules sterility is notrequired. The USP/NF standard is usually sufficient.

In certain embodiments, solid oral dosage forms can further include oneor more components that chemically and/or structurally predispose thecomposition for delivery of the chemical entity to the stomach or thelower GI; e.g., the ascending colon and/or transverse colon and/ordistal colon and/or small bowel. Exemplary formulation techniques aredescribed in, e.g., Filipski, K. J., et al., Current Topics in MedicinalChemistry, 2013, 13, 776-802, which is incorporated herein by referencein its entirety.

Examples include upper-GI targeting techniques, e.g., Accordion Pill(Intec Pharma), floating capsules, and materials capable of adhering tomucosal walls.

Other examples include lower-GI targeting techniques. For targetingvarious regions in the intestinal tract, several enteric/pH-responsivecoatings and excipients are available. These materials are typicallypolymers that are designed to dissolve or erode at specific pH ranges,selected based upon the GI region of desired drug release. Thesematerials also function to protect acid labile drugs from gastric fluidor limit exposure in cases where the active ingredient may be irritatingto the upper GI (e.g., hydroxypropyl methylcellulose phthalate series,Coateric (polyvinyl acetate phthalate), cellulose acetate phthalate,hydroxypropyl methylcellulose acetate succinate, Eudragit series(methacrylic acid-methyl methacrylate copolymers), and Marcoat). Othertechniques include dosage forms that respond to local flora in the GItract, Pressure-controlled colon delivery capsule, and Pulsincap.

Liquid dosage forms for oral administration include pharmaceuticallyacceptable emulsions, microemulsions, solutions, suspensions, syrups andelixirs. In addition to the chemical entities described herein, theliquid dosage forms may contain inert diluents commonly used in the artsuch as, for example, water or other solvents, solubilizing agents andemulsifiers such as ethyl alcohol, isopropyl alcohol, ethyl carbonate,ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol,1,3-butylene glycol, dimethylformamide, oils (in particular, cottonseed,groundnut, corn, germ, olive, castor, and sesame oils), glycerol,tetrahydrofurfuryl alcohol, polyethylene glycols and fatty acid estersof sorbitan, and mixtures thereof. Besides inert diluents, the oralcompositions can also include adjuvants such as wetting agents,emulsifying and suspending agents, sweetening, flavoring, and perfumingagents. In certain embodiments, the liquid dosage form is a mouthwash.In certain embodiments, such liquid oral dosage forms are useful forlocal and topical administration to the digestive or GI tract, e.g.,digestive tract, e.g., oral cavity.

Other Forms of Delivery

In some embodiments, the chemical entities described herein or apharmaceutical composition thereof are suitable for local and topicaladministration to the eye (e.g., eye drops). Ocular compositions caninclude, without limitation, one or more of any of the following:viscogens (e.g., Carboxymethylcellulose, Glycerin, Polyvinylpyrrolidone,Polyethylene glycol); Stabilizers (e.g., Pluronic (triblock copolymers),Cyclodextrins); Preservatives (e.g., Benzalkonium chloride, ETDA, SofZia(boric acid, propylene glycol, sorbitol, and zinc chloride; AlconLaboratories, Inc.), Purite (stabilized oxychloro complex; Allergan,Inc.)).

In some embodiments, the chemical entities described herein or apharmaceutical composition thereof are suitable for local and topicaladministration to skin (e.g., ointments and creams). Ointments aresemisolid preparations that are typically based on petrolatum or otherpetroleum derivatives. Creams containing the selected active agent aretypically viscous liquid or semisolid emulsions, often eitheroil-in-water or water-in-oil. Cream bases are typically water-washable,and contain an oil phase, an emulsifier and an aqueous phase. The oilphase, also sometimes called the “internal” phase, is generallycomprised of petrolatum and a fatty alcohol such as cetyl or stearylalcohol; the aqueous phase usually, although not necessarily, exceedsthe oil phase in volume, and generally contains a humectant. Theemulsifier in a cream formulation is generally a nonionic, anionic,cationic or amphoteric surfactant. As with other carriers or vehicles,an ointment base should be inert, stable, nonirritating andnon-sensitizing.

Dosages

The dosages may be varied depending on the requirement of the patient,the severity of the condition being treating and the particular compoundbeing employed.

Determination of the proper dosage for a particular situation can bedetermined by one skilled in the medical arts. The total daily dosagemay be divided and administered in portions throughout the day or bymeans providing continuous delivery.

In some embodiments, a chemical entity (e.g., a compound exhibitingactivity as a mitochondrial uncoupling agent or a pharmaceuticallyacceptable salt and/or hydrate and/or cocrystal thereof; e.g., acompound, such as niclosamide or a pharmaceutically acceptable saltand/or hydrate and/or cocrystal thereof; e.g., a compound, such as aniclosamide analog, or a pharmaceutically acceptable salt and/or hydrateand/or cocrystal thereof) is administered is administered at a dosage offrom about 0.01 mg/Kg to about 200 mg/Kg (e.g., from about 0.01 mg/Kg toabout 150 mg/Kg; from about 0.01 mg/Kg to about 100 mg/Kg; from about0.01 mg/Kg to about 50 mg/Kg from about 0.01 mg/Kg to about 10 mg/Kgfrom about 0.01 mg/Kg to about 5 mg/Kg; from about 0.1 mg/Kg to about200 mg/Kg; from about 0.1 mg/Kg to about 150 mg/Kg from about 0.1 mg/Kgto about 100 mg/Kg; from about 0.1 mg/Kg to about 50 mg/Kg; from about0.1 mg/Kg to about 10 mg/Kg; from about 0.1 mg/Kg to about 5 mg/Kg).

In certain embodiments, the chemical entity is administered at a dosageof from about 15 mg/Kg to about 100 mg/Kg (e.g., from about 15 mg/Kg toabout 90 mg/Kg, from about 20 mg/Kg to about 100 mg/Kg from about 20mg/Kg to about 90 mg/Kg from about 20 mg/Kg to about 80 mg/Kg from about30 mg/Kg to about 90 mg/Kg; from about 30 mg/Kg to about 80 mg/Kg fromabout 35 mg/Kg to about 75 mg/Kg; from about 10 mg/Kg to about 50 mg/Kg;from about 15 mg/Kg to about 45 mg/Kg; e.g., about 35 mg/Kg or about 75mg/Kg). In other embodiments, the chemical entity is administered at adosage of from about 0.1 mg/Kg to about 10 mg/Kg (e.g., from about 0.1mg/Kg to about 5 mg/Kg; from about 1 mg/Kg to about 10 mg/Kg; from about1 mg/Kg to about 5 mg/Kg).

In some embodiments, enema formulations include from about 0.5 mg toabout 2500 mg (e.g., from about 0.5 mg to about 2000 mg, from about 0.5mg to about 1000 mg, from about 0.5 mg to about 750 mg, from about 0.5mg to about 600 mg, from about 0.5 mg to about 500 mg, from about 0.5 mgto about 400 mg, from about 0.5 mg to about 300 mg, from about 0.5 mg toabout 200 mg; e.g., from about 5 mg to about 2500 mg, from about 5 mg toabout 2000 mg, from about 5 mg to about 1000 mg; from about 5 mg toabout 750 mg; from about 5 mg to about 600 mg; from about 5 mg to about500 mg; from about 5 mg to about 400 mg; from about 5 mg to about 300mg; from about 5 mg to about 200 mg; e.g., from about 50 mg to about2000 mg, from about 50 mg to about 1000 mg, from about 50 mg to about750 mg, from about 50 mg to about 600 mg, from about 50 mg about 500 mg,from about 50 mg to about 400 mg, from about 50 mg to about 300 mg, fromabout 50 mg to about 200 mg; e.g., from about 100 mg to about 2500 mg,from about 100 mg to about 2000 mg, from about 100 mg to about 1000 mg,from about 100 mg to about 750 mg, from about 100 mg to about 700 mg,from about 100 mg to about 600 mg, from about 100 mg to about 500 mg,from about 100 mg to about 400 mg, from about 100 mg to about 300 mg,from about 100 mg to about 200 mg; e.g., from about 150 mg to about 2500mg, from about 150 mg to about 2000 mg, from about 150 mg to about 1000mg, from about 150 mg to about 750 mg, from about 150 mg to about 700mg, from about 150 mg to about 600 mg, from about 150 mg to about 500mg, from about 150 mg to about 400 mg, from about 150 mg to about 300mg, from about 150 mg to about 200 mg; e.g., from about 150 mg to about500 mg; e.g., from about 300 mg to about 2500 mg, from about 300 mg toabout 2000 mg, from about 300 mg to about 1000 mg, from about 300 mg toabout 750 mg, from about 300 mg to about 700 mg, from about 300 mg toabout 600 mg; e.g., from about 400 mg to about 2500 mg, from about 400mg to about 2000 mg, from about 400 mg to about 1000 mg, from about 400mg to about 750 mg, from about 400 mg to about 700 mg, from about 400 mgto about 600 from about 400 mg to about 500 mg; e.g., 150 mg or 450 mg)of the chemical entity in from about 1 mL to about 3000 mL (e.g., fromabout 1 mL to about 2000 mL, from about 1 mL to about 1000 mL, fromabout 1 mL to about 500 mL, from about 1 mL to about 250 mL, from about1 mL to about 100 mL, from about 10 mL to about 1000 mL, from about 10mL to about 500 mL, from about 10 mL to about 250 mL, from about 10 mLto about 100 mL, from about 30 mL to about 90 mL, from about 40 mL toabout 80 mL; from about 50 mL to about 70 mL; e.g., about 1 mL, about 5mL, about 10 mL, about 15 mL, about 20 mL, about 25 mL, about 30 mL,about 35 mL, about 40 mL, about 45 mL, about 50 mL, about 55 mL, about60 mL, about 65 mL, about 70 mL, about 75 mL, about 100 mL, about 250mL, or about 500 mL, or about 1000 mL, or about 2000 mL, or about 3000mL; e.g., 60 mL) of liquid carrier.

In certain embodiments, enema formulations include from about 50 mg toabout 250 mg (e.g., from about 100 mg to about 200; e.g., about 150 mg)of the chemical entity in from about 10 mL to about 100 mL (e.g., fromabout 20 mL to about 100 mL, from about 30 mL to about 90 mL, from about40 mL to about 80 mL; from about 50 mL to about 70 mL) of liquidcarrier. In certain embodiments, enema formulations include about 150 mgof the chemical entity in about 60 mL of the liquid carrier. In certainof these embodiments, the chemical entity is niclosamide, or apharmaceutically acceptable salt and/or hydrate and/or cocrystalthereof. For example, enema formulations can include about 150 mg ofniclosamide in about 60 mL of the liquid carrier.

In certain embodiments, enema formulations include from about 350 mg toabout 550 mg (e.g., from about 400 mg to about 500; e.g., about 450 mg)of the chemical entity in from about 10 mL to about 100 mL (e.g., fromabout 20 mL to about 100 mL, from about 30 mL to about 90 mL, from about40 mL to about 80 mL; from about 50 mL to about 70 mL) of liquidcarrier. In certain embodiments, enema formulations include about 450 mgof the chemical entity in about 60 mL of the liquid carrier. In certainof these embodiments, the chemical entity is niclosamide, or apharmaceutically acceptable salt and/or hydrate and/or cocrystalthereof. For example, enema formulations can include about 450 mg ofniclosamide in about 60 mL of the liquid carrier.

In some embodiments, enema formulations include from about from about0.01 mg/mL to about 50 mg/mL (e.g., from about 0.01 mg/mL to about 25mg/mL; from about 0.01 mg/mL to about 10 mg/mL; from about 0.01 mg/mL toabout 5 mg/mL; from about 0.1 mg/mL to about 50 mg/mL; from about 0.01mg/mL to about 25 mg/mL; from about 0.1 mg/mL to about 10 mg/mL; fromabout 0.1 mg/mL to about 5 mg/mL; from about 1 mg/mL to about 10 mg/mL;from about 1 mg/mL to about 5 mg/mL; from about 5 mg/mL to about 10mg/mL; e.g., about 2.5 mg/mL or about 7.5 mg/mL) of the chemical entityin liquid carrier. In certain of these embodiments, the chemical entityis niclosamide, or a pharmaceutically acceptable salt and/or hydrateand/or cocrystal thereof. For example, enema formulations can includeabout 2.5 mg/mL or about 7.5 mg/mL of niclosamide in liquid carrier.

The foregoing dosages can be administered on a daily basis (e.g., as asingle dose per day; or as two or more divided doses per day; or a twoor more doses; e.g., two doses per day) or non-daily basis (e.g., everyother day, every two days, every three days, once weekly, twice weeks,once every two weeks, once a month). In certain embodiments, dosages canbe administered for about 1 week, about 2 weeks, about 3 weeks, about 4weeks, about 5 weeks, about 6 weeks, about 7 weeks, about 8 weeks, about3 months, about 6 months, about 1 year, or beyond. For example, dosages(e.g., about 2.5 mg/mL or about 7.5 mg/mL) of the chemical entity inliquid carrier can be administered twice a day on a daily basis forabout 6 weeks. In certain of these embodiments, the chemical entity isniclosamide, or a pharmaceutically acceptable salt and/or hydrate and/orcocrystal thereof. For example, about 2.5 mg/mL or about 7.5 mg/mL ofniclosamide in liquid carrier can be administered twice a day on a dailybasis for about 6 weeks. Representative liquid carriers include, e.g.,those previously described in conjunction with component (ii).

Methods of Treatment

In some embodiments, methods for inducing cell death of one or more Tcells (e.g., in the digestive and/or gastrointestinal tract (GI), skin,eyes, or joints), of a subject are provided. The methods includecontacting the one or more T cells with an effective amount of achemical entity (e.g., a compound exhibiting activity as a mitochondrialuncoupling agent or a pharmaceutically acceptable salt and/or hydrateand/or cocrystal thereof; e.g., a compound, such as niclosamide or apharmaceutically acceptable salt and/or hydrate and/or cocrystalthereof; e.g., a compound, such as a niclosamide analog, or apharmaceutically acceptable salt and/or hydrate and/or cocrystalthereof) as defined anywhere herein. In certain embodiments, the methodsconsist essentially or consist of the contacting step described above inthis paragraph.

In some embodiments, methods for treating a subject having a conditionassociated with unregulated (abnormal, elevated) recruitment and/orretention of one or more T cells (e.g., at the digestive and/orgastrointestinal tract (GI), skin, eyes, or joints) of the subject areprovided. The methods include contacting the one or more T cells with aneffective amount of a chemical entity (e.g., a compound exhibitingactivity as a mitochondrial uncoupling agent or a pharmaceuticallyacceptable salt and/or hydrate and/or cocrystal thereof, e.g., acompound, such as niclosamide or a pharmaceutically acceptable saltand/or hydrate and/or cocrystal thereof; e.g., a compound, such as aniclosamide analog, or a pharmaceutically acceptable salt and/or hydrateand/or cocrystal thereof) as defined anywhere herein. In certainembodiments, the methods consist essentially of or consist of thecontacting step described above in this paragraph.

In some embodiments, methods for treating a subject having a conditionassociated with unregulated (abnormal, elevated) activation of one ormore T cells (e.g., in the digestive and/or gastrointestinal tract (GI),skin, eyes, or joints) of the subject are provided. The methods includecontacting the one or more activated T cells with an effective amount ofa cocrystal comprising (i) a mitochondrial uncoupling agent or apharmaceutically acceptable salt and/or hydrate thereof; and (ii) one ormore pharmaceutically acceptable coformers as defined anywhere herein.In certain embodiments, the methods consist essentially of or consist ofthe contacting step described above in this paragraph.

In some embodiments, inducing cell death of the one or more T cellsincludes one or more of the following pathways: Programmed cell death,Necroptosis, Apoptosis, Necrosis, Pyroptosis, Ferroptosis, Anoikis,Mitotic cathastrophe, Paraptosis, Pyronecrosis, Entosis, Netosis,Parthanatos, Autophagic cell death, RGD: regulated cell death,Non-apoptotic programmed cell-death, Caspase-independent programmedcell-death inducing necrosis or apoptosis of the one or more T cells,e.g., necrosis or apoptosis of the one or more T cells. In certainembodiments, the effective amount is an amount sufficient to induce celldeath of at least one of the one or more T cells (e.g., by any one ormore of the pathways described above, e.g., necrosis or apoptosis of theone or more T cells).

In some embodiments, the one or more T cells include one or moreactivated T cells, e.g., one or more activated T cells is independentlyselected from the group consisting of:

CD45+CD3+TCRαβ+CD62L−;

CD45+CD3+TCRαβ+CD62L−CCR7−;

CD45+CD3+TCRαβ+CD62L−CD69+;

CD45+CD3+TCRαβ+CD62L−CD69+PD−1+;

CD45+CD3+TCRαβ+CD62L−CTLA4+;

CD45+CD3+TCRαβ+CD62L−PD−1++CTLA4+;

CD45+CD3+TCRγδ+CD62L−;

CD45+CD3+TCRγδ+CD62L−CCR7−;

CD45+CD3+TCRγδ+CD62L−CD69+;

CD45+CD3+TCRγδ+CD62L−CD69+PD−1+;

CD45+CD3+CD62L−TCRγδ+CTLA4+; and

CD45+CD3+TCRγδ+CD62L−PD−++CTLA4+.

In certain embodiments, the effective amount is an amount sufficient toinduce cell death of at least one of the one or more activated T cells(e.g., by any one or more of the pathways described above, e.g.,necrosis or apoptosis of the one or more activated T cells).

In some embodiments, the one or more T cells are present within theintestinal epithelium and/or within the lamina propria and/or within thePeyer's patches (PP) and/or within the GALT (gut associated lymphoidtissue) and/or within the intestinal mucosa and/or within the intestinalsubmucosa and/or within the intestinal muscular layer and/or within theintestinal serosa.

In some embodiments, the one or more T cells comprise one or more guttropic T cells. In certain embodiments, each of the one or more guttropic T cells independently expresses one or more gut-homing receptorsselected from the group consisting of:

(CD3+CCR9+;

CD3+α4+ or CD3+β7+;

CD3+α4+β7+;

CD3+β+;

CD3+α4+β1+;

CD3+LFA1;

CD3+CCR4+; and

CD3+CCR10+.

In some embodiments, methods for treating a condition (or one or moresymptoms thereof) characterized by an abnormal inflammatory response ina subject in need thereof are provided (e.g., an autoimmune disorder,e.g., an inflammatory bowel disease). The methods include administeringto the subject an effective amount of a chemical entity (e.g., acompound exhibiting activity as a mitochondrial uncoupling agent or apharmaceutically acceptable salt and/or hydrate and/or cocrystalthereof; e.g., a compound, such as niclosamide or a pharmaceuticallyacceptable salt and/or hydrate and/or cocrystal thereof; e.g., acompound, such as a niclosamide analog, or a pharmaceutically acceptablesalt and/or hydrate and/or cocrystal thereof) as defined anywhereherein. In certain embodiments, the methods consist essentially of orconsist of the administering step described above in this paragraph.

In some embodiments, methods for treating a condition (or one or moresymptoms thereof) characterized by an abnormal inflammatory response ina subject in need thereof are provided (e.g., an autoimmune disorder,e.g., an inflammatory bowel disease). The methods include topically andlocally administering to the subject an effective amount of a chemicalentity (e.g., a compound exhibiting activity as a mitochondrialuncoupling agent or a pharmaceutically acceptable salt and/or hydrateand/or cocrystal thereof; e.g., a compound, such as niclosamide or apharmaceutically acceptable salt and/or hydrate and/or cocrystalthereof; e.g., a compound, such as a niclosamide analog, or apharmaceutically acceptable salt and/or hydrate and/or cocrystalthereof) as defined anywhere herein. In certain embodiments, the methodsconsist essentially of or consist of the administering step describedabove in this paragraph.

In some embodiments, methods for treating autoimmune colitis (or one ormore symptoms thereof) in a subject are provided. The methods includetopically and locally administering to the subject an effective amountof a chemical entity (e.g., a compound exhibiting activity as amitochondrial uncoupling agent or a pharmaceutically acceptable saltand/or hydrate and/or cocrystal thereof; e.g., a compound, such asniclosamide or a pharmaceutically acceptable salt and/or hydrate and/orcocrystal thereof; e.g., a compound, such as a niclosamide analog, or apharmaceutically acceptable salt and/or hydrate and/or cocrystalthereof) as defined anywhere herein. In certain embodiments, the methodsconsist essentially of or consist of the administering step describedabove in this paragraph.

In some embodiments, methods for treating a condition (or one or moresymptoms thereof) selected from the group consisting of celiac disease,irritable bowel syndrome, mucositis, uveitis, collagenous colitis,lymphocytic colitis, microscopic colitis, radiation enteritis,rheumatoid arthritis, lupus, scleroderma, psoriasis, cutaneous T-celllymphoma, acute graft vs. host disease and chronic graft vs. hostdisease in a subject are provided. The methods include topically andlocally administering to the subject an effective amount of a chemicalentity (e.g., a compound exhibiting activity as a mitochondrialuncoupling agent or a pharmaceutically acceptable salt and/or hydrateand/or cocrystal thereof; e.g., a compound, such as niclosamide or apharmaceutically acceptable salt and/or hydrate and/or cocrystalthereof; e.g., a compound, such as a niclosamide analog, or apharmaceutically acceptable salt and/or hydrate and/or cocrystalthereof) as defined anywhere herein. In certain embodiments, the methodsconsist essentially of or consist of the administering step describedabove in this paragraph.

In certain of these embodiments, the condition is an autoimmune disease.In certain embodiments, the condition is an inflammatory bowel disease.In certain embodiments, the condition is Crohn's disease, autoimmunecolitis, iatrogenic autoimmune colitis, ulcerative colitis, colitisinduced by one or more chemotherapeutic agents, colitis induced bytreatment with adoptive cell therapy, colitis associated by one or morealloimmune diseases (such as graft-vs-host disease, e.g., acute graftvs. host disease and chronic graft vs. host disease), radiationenteritis, collagenous colitis, lymphocytic colitis, microscopiccolitis, and radiation enteritis.

In certain of these embodiments, the condition is alloimmune disease(such as graft-vs-host disease, e.g., acute graft vs. host disease andchronic graft vs. host disease), celiac disease, irritable bowelsyndrome, rheumatoid arthritis, lupus, scleroderma, psoriasis, cutaneousT-cell lymphoma, uveitis, and mucositis (e.g., oral mucositis,esophageal mucositis or intestinal mucositis).

In certain embodiments, the condition is autoimmune colitis.

In certain of these embodiments, the autoimmune colitis is induced byone or more chemotherapeutic agents, e.g., a chemotherapeuticimmunomodulator, e.g., an immune checkpoint inhibitor. In certain ofthese embodiments, the immune checkpoint inhibitor targets an immunecheckpoint receptor selected from the group consisting of CTLA-4, PD-1,PD-L1, PD-1-PD-L1, PD-1-PD-L2, interleukin-2 (IL-2), indoleamine2,3-dioxygenase (IDO), IL-10, transforming growth factor-β (TGFβ), Tcell immunoglobulin and mucin 3 (TIM3 or HAVCR2), Galectin 9-TIM3,Phosphatidylserine-TIM3, lymphocyte activation gene 3 protein (LAG3),MHC class II-LAG3, 4-BB-4-BB ligand, OX40-OX40 ligand, GITR, GITRligand-GITR, CD27, CD70-CD27, TNFRSF25, TNFRSF25-TLA, CD40L, CD40-CD40ligand, HVEM-LIGHT-LTA, HVEM, HVEM-BTLA, HVEM-CD160, HVEM-LIGHT,HVEM-BTLA-CD160, CD80, CD80-PDL-1, PDL2-CD80, CD244, CD48-CD244, CD244,ICOS, ICOS-ICOS ligand, B7-H3, B7-H4, VISTA, TMIGD2, HHLA2-TMIGD2,Butyrophilins, including BTNL2, Siglec family, TIGIT and PVR familymembers, KIRs, ILTs and LIRs, NKG2D and NKG2A, MICA and MICB, CD244,CD28, CD86-CD28, CD86-CTLA, CD80-CD28, CD39, CD73 Adenosine-CD39-CD73,CXCR4-CXCL12, Phosphatidylserine, TIM3, Phosphatidylserine-TIM3,SIRPA-CD47, VEGF, Neuropilin, CD160, CD30, and CD155; e.g., CTLA-4 orPD1 or PD-L1). See, e.g., Postow, M. J. Clin. Oncol. 2015, 33, 1.

In certain of these embodiments, the immune checkpoint inhibitor isselected from the group consisting of: Urelumab, PF-05082566, MEDI6469,TRX518, Varlilumab, CP-870893, Pembrolizumab (PD1), Nivolumab (PD1),Atezolizumab (formerly MPDL3280A) (PDL), MEDI4736 (PD-L1), Avelumab(PD-L1), PDR001 (PD1), BMS-986016, MGA271, Lirilumab, IPH2201,Emactuzumab, INCB024360, Galunisertib, Ulocuplumab, BKT140, Bavituximab,CC-90002, Bevacizumab, and MNRP1685A, and MGA271.

In certain of these embodiments, the immune checkpoint inhibitor targetsCTLA-4, e.g., an antibody, e.g., ipilimumab or tremelimumab.

In certain of these embodiments, the immune checkpoint inhibitor targetsPD1 or PD-L1, e.g., nivolumab, lambroizumab, or BMS-936559.

In certain embodiments, the condition is mucositis, also known asstomatitits, which can occur as a result of chemotherapy or radiationtherapy, either alone or in combination as well as damage caused byexposure to radiation outside of the context of radiation therapy.Chemotherapeutic agents which may induce mucositis when used alone or incombination include, but are not limited to, platinum, cisplatin,carboplatin, oxaliplatin, mechlorethamine, cyclophosphamide,chlorambucil, azathioprine, mercaptopurine, vincristine, vinblastine,vinorelbine, vindesine, etoposide and teniposide, paclitaxel, docetaxel,irinotecan, topotecan, amsacrine, etoposide, etoposide phosphate,teniposide, 5-fluorouracil, leucovorin, methotrexate, gemcitabine,taxane, leucovorin, mitomycin C, tegafur-uracil, idarubicin,fludarabine, mitoxantrone, ifosfamide and doxorubicin. Additional agentsinclude inhibitors of mTOR (mammalian target of rapamycin), includingbut not limited to rapamycin, everolimus, temsirolimus and deforolimus.

In certain embodiments, the condition is uveitis, which is inflammationof the uvea (e.g., anterior uveitis, e.g., iridocyclitis or iritis;intermediate uveitis (also known as pars planitis); posterior uveitis;or chorioretinitis, e.g., pan-uveitis).

This disclosure contemplates both monotherapy regimens as well ascombination therapy regimens.

In some embodiments, monotherapy includes administering (e.g., topicallyand locally) to a subject an effective amount of a chemical entity(e.g., a compound exhibiting activity as a mitochondrial uncouplingagent or a pharmaceutically acceptable salt and/or hydrate and/orcocrystal thereof; e.g., a compound, such as niclosamide or apharmaceutically acceptable salt and/or hydrate and/or cocrystalthereof; e.g., a compound, such as a niclosamide analog, or apharmaceutically acceptable salt and/or hydrate and/or cocrystalthereof) as defined anywhere herein, but excludes the administration ofother therapeutic agents (e.g., the active compounds, e.g., peptides,disclosed in U.S. Pat. No. 8,148,328, which is incorporated herein byreference in its entirety).

In some embodiments, the methods described herein can further includeadministering a second therapeutic agent or regimen.

In certain embodiments, the second therapeutic agent or regimen isadministered to the subject prior to contacting with or administeringthe chemical entity (e.g., about one hour prior, or about 6 hours prior,or about 12 hours prior, or about 24 hours prior, or about 48 hoursprior, or about 1 week prior, or about 1 month prior).

In other embodiments, the second therapeutic agent or regimen isadministered to the subject at about the same time as contacting with oradministering the chemical entity. By way of example, the secondtherapeutic agent or regimen and the chemical entity are provided to thesubject simultaneously in the same dosage form. As another example, thesecond therapeutic agent or regimen and the chemical entity are providedto the subject concurrently in separate dosage forms.

In still other embodiments, the second therapeutic agent or regimen isadministered to the subject after contacting with or administering thechemical entity (e.g., about one hour after, or about 6 hours after, orabout 12 hours after, or about 24 hours after, or about 48 hours after,or about 1 week after, or about 1 month after).

In certain embodiments, the second therapeutic agent is achemotherapeutic immunomodulator, e.g., an immune checkpoint inhibitor,which can be as defined anywhere herein. In other embodiments, thesecond therapeutic agent or regimen is one or more anti-inflammatoryagents or immunomodulator acting locally in the GI tract. In otherembodiments, the second therapeutic agent or regimen is 5-ASA (andassociated delivery systems), anti-SMAD7 antisense, orally formulatedanti-TNFs, anti-integrins, sulfasalazine, balsalazide, steroids,azathioprine, and methotrexate. In further embodiments, the secondtherapeutic agent or regimen is radiation or surgery.

In certain embodiments, the second therapeutic agent is platinum,cisplatin, carboplatin, oxaliplatin, mechlorethamine, cyclophosphamide,chlorambucil, azathioprine, mercaptopurine, vincristine, vinblastine,vinorelbine, vindesine, etoposide and teniposide, paclitaxel, docetaxel,irinotecan, topotecan, amsacrine, etoposide, etoposide phosphate,teniposide, 5-fluorouracil, leucovorin, methotrexate, gemcitabine,taxane, leucovorin, mitomycin C, tegafur-uracil, idarubicin,fludarabine, mitoxantrone, ifosfamide and doxorubicin. Additional agentsinclude inhibitors of mTOR (mammalian target of rapamycin), includingbut not limited to rapamycin, everolimus, temsirolimus and deforolimus.

In still other embodiments, the second therapeutic agent can be selectedfrom those delineated above (see U.S. Pat. No. 7,927,613, which isincorporated herein by reference in its entirety).

In some embodiments, the methods described herein further include thestep of identifying a subject (e.g., a patient) in need of suchtreatment (e.g., by way of biopsy, endoscopy, or other conventionalmethod known in the art).

In some embodiments, the chemical entities, methods, and compositionsdescribed herein can be administered to certain treatment-resistantpatient populations, e.g., one that is nonresponsive or resistant totreatment with an anti-TNFalpha therapy (e.g., Humira, Enbrel, Remicade,Cimzia, Simponi, Enbrel, xanthine derivatives, e.g., pentoxifylline andBupropion; (R)-DOI, TCB-2, LSD and LA-SS-Az). In certain embodiments,the patient is undergoing and/or has undergone treatment with ananti-TNFalpha therapy (e.g., Humira, Enbrel, Remicade, Cimzia, Simponi,Enbrel, xanthine derivatives, e.g., pentoxifylline and Bupropion;(R)-DOI, TCB-2, LSD and LA-SS-Az).

To further illustrate this invention, the following examples areincluded. The examples should not, of course, be construed asspecifically limiting the invention. Variations of these examples withinthe scope of the claims are within the purview of one skilled in the artand are considered to fall within the scope of the invention asdescribed, and claimed herein. The reader will recognize that theskilled artisan, armed with the present disclosure, and skill in the artis able to prepare and use the invention without exhaustive examples.

EXAMPLES Example 1: Niclosamide Uncouples Mitochondrial Respiration fromOxidative Phosphorylation Jurkat T Cells

Objective.

To measure the dose-response effect of niclosamide on mitochondrialtransmembrane potential in Jurkat T cells using the lipophilic cationicdye, tetramethylrhodamine, methyl ester (TMRM).

Model.

The Jurkat T cell model is commonly used to study the potential effectsof compounds on T cells in vitro. This cell line allows investigation ofstimuli and mechanisms that regulate T cell mitochondrial function andsurvival. As T cells, Jurkats have a lymphocyte appearance and replicatein culture in suspension. They contain respiring mitochondria and assuch response to mitochondrial uncouplers such as niclosamide may beassessed. Uncoupling is identified and quantified by a detecting a dropin the electrochemical gradient across the mitochondrial inner membrane(ΔΨm) that is not associated with a corresponding increase in oxidativephosphorylation. Experiments to detect changes in ΔΨm were performed byincluding conditions in which a concentration of oligomycin was added toirreversibly inhibit the F₁F₀-ATPase and block oxidative phosphorylationto demonstrate that the fall in ΔΨm represents uncoupling since itoccurred independent of an increase in mitochondrial oxidativephosphorylation.

Cell Culture.

Jurkat T cells were purchased from the American Type Culture Collection(Manassas, Va.) and sub-cultured according to instructions from thesupplier. Prior to experiments, cells were cultured in RPMI 1640containing 100% FBS-HI, 50 units penicillin/mL and 50 μgstreptomycin/mL, and maintained in log phase prior to experimentalsetup. Cells were grown in a 5% CO₂ humidified incubator at 37° C.Growth medium was made by adding 50 mL of heat inactivated FBS and 5 mLof penicillin/streptomycin to 500 mL DMEM. This medium was stored at 4°C. Before use, the medium was warmed to 37° C. in a water bath. Jurkatcells were seeded at an initial density of 5×10⁴ cells/mL in 24-wellplates. The cells were allowed to grow for 18 hours prior to treatmentbeing added.

Treatment with Niclosamide.

Niclosamide was dissolved in dimethyl sulfoxide (DMSO) and added to theculture medium to achieve concentrations of 500, 100, 50, 10, 5 or 1 μM.Oligomycin was dissolved in DMSO then added to test wells in 10 μL toachieve a final concentration of 1 μL. Samples were incubated for 60minutes at 37° C. TMRM dissolved in DMSO then added to the test wells in10 μL to achieve a final concentration of 5 μM and allowed to incubateat 37° C. for an additional 30 min. A vehicle only control (in place ofniclosamide) was run concurrently with each experiment. A flow cytometerproviding excitation at 560 nm and detection at 590 nm emission was usedfor quantification of TMRM fluorescence.

Measuring Mitochondrial Membrane Potential Changes (ΔΨm).

TMRM has advantages over other cationic dyes in that it can selectivelyenter into mitochondria and reversibly accumulate as the membranepotential increases. The accumulation of TMRM in mitochondria has beenshown to be driven by their membrane potential Moreover, because ofreduced hydrophobic character, this probe exhibits potential-independentbinding to cells that is 10 to 20 times lower than that seen with otherprobes. TMRM has been described as one of the best fluorescent dyes fordynamic and in situ quantitative measurements because it is rapidly andreversibly taken up by live cells and mitochondria.

Calculation of Relative Decrease in Mitochondrial Membrane Potential.

Median fluorescence intensity was computed for all concentrations ofniclosamide relative to vehicle-only negative controls in the presenceof oligomycin. Ratios of the fluorescence intensity of each treatedsample to the control sample mean were then calculated as a measure ofrelative decrease in ΔΨm. For statistical comparisons, 95% confidenceintervals were computed and graphed with the mean values of this ratio.By utilizing the 95% confidence intervals, the probability of a type Ierror was set at the nominal 5% level.

Results.

Niclosamide exhibits a dose-related decrease in ΔΨm in Jurkat cells withconcentrations of niclosamide of 5 μM and above significantly decreased(p<0.05) relative to negative controls.

Example 2: Niclosamide Uncouples Mitochondrial Respiration fromOxidative Phosphorylation in T Cells Isolated from the Lamina Propria ofHuman Intestine

Objective.

The objective of this experiment is to determine if niclosamide candirectly reduce the mitochondrial transmembrane potential in T cellsisolated from human intestine lamina propria in a manner similar toeffects observed in Jurkat T cells.

Model.

Lamina propria mononuclear cells (LPMC) in the human intestine arecomprised in part by T cells, which mediate physiological andpathological processes including inflammatory bowel disease. LPMCs canbe isolated from human tissue biopsies. After isolation LPMCs T cellsremain viable ex vivo under appropriate culture conditions for periodsof time that allow ex vivo experiments. These cells can be used toinvestigate mechanisms that regulate their mitochondrial function andsurvival. They contain respiring mitochondria and as such their responseto mitochondrial uncouplers such as niclosamide may be assessed. Thiscellular model is used in conjunction with oligomycin that blocksoxidative phosphorylation and TMRM to monitor ΔΨm as described inExample 1.

Cell Isolation and Culture.

Cells are obtained from biopsy specimens of the small or large intestineor rectum of humans from areas of normal gastrointestinal tissue or withmoderate to severe Crohn's disease (CD), ulcerative colitis (UC), orceliac disease. For the isolation of lamina propria mononuclear cells(LPMCs), the specimens are initially washed in Hank's balanced saltsolution (HBSS) then are cut into 0.5-cm pieces, and are incubated withstirring in pre-warmed HBSS containing 1 mM DTT at 37° C. for 15minutes. The supernatant is removed and the sample is washed withstirring with HBSS for 5 minutes twice. Samples are incubated withstirring in pre-warmed HBSS containing 5 mM EDTA for 30 minutes. Thesupernatant is removed and the sample is washed with stirring with HBSSfor 5 minutes three times. The tissue is then digested further in RPMI1640 containing 2 mg/mL Liberase and 0.01 μg/mL DNase I for 1 hour at37° C. with stirring. After digestion, the mononuclear cells insuspension are collected and are centrifuged at 400 g for 10 minutes.After two washings in HBS, the pellet is resuspended in a 40% Percollsolution and is layered on the top of a Percoll solution (100%, 60%,40%, and 30% Percoll in HBSS.). The tube is centrifuged at 400 g for 25minutes, and LPMCs at the 60%-40% Percoll layer interface are collected.The isolated cells are counted and checked for viability using 0.1%trypan blue (viability ranges from 86% to 94%). Cells are washed out ofPercoll with HBSS and are resuspended in RPMI 1640 supplemented with 10%heat inactivated FBS, 1% L-glutamine, 100 U/mL penicillin, and 100 mg/mLstreptomycin at a concentration of 1×10⁶ cells/mL and are plated in96-well culture plates (200000 cells/well) (Nat Protoc. 2007;2(10):2307-11).

Treatment with Test Material.

The protocol as noted in Example 1 is followed. In addition, anti-CD3monoclonal antibody conjugated to FITC (excitation at 494 nm withemission detected at 521 nm) is additionally added during incubationwith TMRM at 37° C. for 30 minutes.

Measurement of and Calculation of Change of ΔΨm in T Cells.

In order to specifically distinguish T cells from other cells in LPMCs,anti-CD3 monoclonal antibody labeled with FITC is used. This antibodyspecifically binds human CD3 antigen that is selectively expressed on Tcells. LPMC T cells are first defined by their fluorescence emission at521 nm resulting from labeling with FITC-anti-CD3 antibody. The nfluorescence intensity of TMRM detected at 590 nm in the T cellpopulation is measured. Median fluorescence intensity of the TMRM signalare computed. Ratios of the median fluorescence intensity of eachtreated sample to the control sample mean are then calculated as ameasure of relative decrease in ΔΨm. For statistical comparisons, 95%confidence intervals are computed and graphed with the mean values ofthis ratio.

Results.

Niclosamide induces a dose-related decrease in ΔΨm in human LPMCs Tcells with concentrations of niclosamide of 5 μM and above (p<0.05)relative to negative control s.

Example 3: Niclosamide Induces Death of LPMC T Cells at Concentrationsthat Cause Mitochondrial Uncoupling

Objective.

The objective of this experiment was to determine if concentrations ofniclosamide that uncouple mitochondria in LPMC induce cell death.

Model.

The human LPMC model as described in Example 2 was used.

Cell Isolation and Culture.

Cell isolation and culture procedures were as detailed in Example 2.

Treatment with Niclosamide.

Niclosamide was dissolved in dimethyl sulfoxide (DMSO) and added to theculture medium to achieve concentrations of 500, 100, 50, 10, 5 or 1 μM.Samples were incubated for 60 minutes at 37° C. Cultured cells wereincubated with DMSO (negative control), or stimulated with a humanmonoclonal anti-FAS-activating antibody (positive control, finalconcentration, 1 μg/mL), or concentrations of niclosamide at 37° C. for24 hours. After treatment cells were exposed to 1 μM 7AAD then incubateda further 60 minutes at 37° C. Live cells and dead cells were enumeratedby flow cytometer using a FACSVerse cytometer set to excite and measureemitted fluorescence of 7-AAD at appropriate wavelengths.

Detecting Viable and Dead Cells.

7-AAD is excluded from live cells but free to enter dead cells where itundergoes a spectral shift after interacting with cellular DNA. Thusdead cells are selectively labeled with 7-AAD resulting in theirdetection with an emission maxima of 647 nm. Use of this reagent allowsviable cells and dead cells to be simultaneously enumerated. In order tospecifically distinguish T cells from other cells in LPMCs, anti-CD3monoclonal antibody labeled with FITC (excitation at 494 nm withemission detected at 521 nm) will be used. This antibody specificallybinds human CD3 antigen that is selectively expressed on T cells. Cellviability and death was determined specifically in T cells by measuring7-AAD fluorescence in cells labeled by anti-CD3 FITC.

Calculation of T Cell Death.

The fluorescence intensity of 7-AAD detected at 647 nm was measuredspecifically T the cell population that was first defined as describedin Example 2 by FITC-anti-CD3 antibody fluorescence. In each experimentthe 7-AAD fluorescence intensity value below which >95% of untreatedcontrol (live) T cells were detected was used as a cut point tocalculate viability. Using this cut-point, the fraction of dead cells ina sample of >10,000 individual cells was calculated for each conditionand expressed as mean values. For statistical comparisons, 95%confidence intervals were computed and graphed with the mean values.

Results.

Niclosamide exhibits a dose-related increase in LPMC T cell death.Concentrations of niclosamide of 5 μM and above significantly increasedeath (p<0.05) relative to negative controls with vehicle alone.Concentrations below 5 μM fail to induce T cell death. The dose-responserelationships of niclosamide-associated T cell death and niclosamideassociated uncoupling in LPMCs were compared. The overlapping nature ofthese dose relationships indicates an association between niclosamideinduced mitochondrial uncoupling and cell death.

Example 4: Niclosamide is an Effective Treatment for Inflammatory BowelDisease in Mice

Objective.

The objective of this experiment is to determine if niclosamide is aneffective treatment in a rodent model of colitis.

Model.

The TNBS-induced colitis is a commonly used experimental model forInflammatory Bowel Disease (IBD). TNBS (trinitrobenzenesulfonic acid) isa chemical administered rectally in the form of an enema to mice or ratsin combination with ethanol, which disrupts the mucous barrier, andinduces colitis by haptenating proteins within the gut, causing them tobecome preferential targets for immune cells. The severity ofTNBS-induced colitis depends largely on the dosage applied and thestrain of animal used. In chronic, relapsing form of the model, animalsare sensitized by escalating, intracolonic doses of TNBS. Disease ismonitored in-life by weight loss. Histology of colon specimens is usedto determine disease severity at study termination (Gastroenterology.2003 December; 125(6): 1750-61; Inflamm Bowel Dis. 2006 October;12(10):995-9.)

Mouse Strain and Housing.

C57BL/6J female mice (9-weeks old) are purchased by Jackson and arehoused at a temperature ranging from 68 to 74° F. with a diurnal 12 hourlight cycle in a specific pathogen-free facility in ventilated cages.Food and water is provided ad libitum. Animals are acclimated to localmicrobiota for 7 days before the beginning of the experiment. Cellisolation and culture procedures are as noted in Example 2.

Conditioning to Induce Colitis.

To perform the studies of relapsing hapten-induced colitis 4 escalatingdoses of TNBS in 50% ethanol are administered at weekly intervals tolightly anesthetized mice through a 3.5 Fr catheter inserted into therectum. The catheter tip is inserted 4 cm proximal to the anal verge,and 150 μL of fluid is slowly instilled into the colon, after which themouse is held in a vertical position for 30 seconds per rectum at weeklyintervals. The first and second doses are 0.5 mg TNBS, whereas the thirdand fourth doses are 0.75 and 1 mg TNBS. A control group is administeredevery week with 50% ethanol using the same procedure. Animal niclosamideis dissolved in water and is administered at 1, 3, 10, 30, 100 mg/kg atdaily intervals to lightly anesthetized mice through a 3.5 Fr catheterinserted into the rectum. Control mice are administered with water usingthe same procedure.

Clinical Assessment of Disease.

For the clinical assessment of colitis, animal weight, diarrhea(0=absent; 1=present), rectal prolapse (0=absent; 1=present) andpresence of blood in the stool (0=absent; 1=present) is recorded daily.

Histological Assessment of Disease.

For histologic analysis, tissues are fixed in OCT, are cut intosections, and are stained with H&E. Histology scoring for individualmice is performed by a pathologist blinded to the samples, and thedegree of inflammation on microscopic cross-sections of the colon isgraded semiquantitatively from 0 to 4. Tissues that are removed frommice at indicated times of death are fixed in 10% formalin solution, areembedded in paraffin, are cut into tissue sections and are stained withhematossiline and eosine. Stained sections are examined for evidence ofcolitis using different criteria such as the presence of lymphocyteinfiltration, elongation and/or distortion of crypts, frank ulcerationand thickening of the bowel wall. The degree of inflammation onmicroscopic cross-sections of the colon is graded from 0 to 4 asfollows: 0: no evidence of inflammation; 1: low level of lymphocyteinfiltration with infiltration seen in a <10% high-power field (hpf=highpower field), no structural changes observed; 2: moderate lymphocyteinfiltration with infiltration seen in <10-25% hpf, crypt elongation,bowel wall thickening which does not extend beyond mucosal layer; 3:high level of lymphocyte infiltration with infiltration seen in <25-50%hpf, thickening of bowel wall which extends beyond mucosal layer; 4:marked degree of lymphocyte infiltration with infiltration seen in >50%hpf, high vascular density, crypt elongation with distortion, transmuralbowel wall-thickening with ulceration (J Exp Med. 1995 Nov. 1;182(5):1281-90; Current Protocol Immunology 15.19 DOI:10.1002/0471142735.im1519s49).

Calculation of Therapeutic Effects.

For statistical comparisons, two-way Anova testwith Bonferronicorrection is calculated with GaphPrism software.

Results.

Niclosamide exhibits a dose-related decrease in colitis clinical scoresand histologic scores. Therapeutic doses of niclosamide of 3 mg/kg andabove significantly reduce both clinical and histologic scores (p<0.05)relative to vehicle control.

Example 5: Therapeutic Doses of Niclosamide in Mice are Associated witha Colon to Plasma Exposure Ratio that Exceeds 10:1

Objective.

The objective of this experiment is to determine plasma and colonconcentration and calculate colon to plasma exposure in mice dosedrectally with niclosamide.

Model.

Mice are used as an effective model to correlate therapeutic responseswith the drug concentrations that can be measured in the blood (serum orplasma fraction) and in tissues to determine the effectiveness of atreatment strategy designed to provide colon topical administration asopposed to systemic absorption. By measuring test agent concentrationsin the strain of mice in which therapeutic responses to colitis areobserved, conclusions can be reached as to whether topical colonicdelivery produces a high colon:plasma ratio of drug exposure andsufficient colon concentrations of the test agent to account fortherapeutic effects that are independent of absorption and systemicexposure to the test agent.

Mice.

Nine week old C57BL/6J female mice are purchased by Jackson and arehoused at a temperature ranging from 68 to 74° F. with a diurnal 12 hourlight cycle in a specific pathogen-free facility in ventilated cages.Food and water is provided ad libitum. Animals are acclimated to localmicrobiota for 7 days before the beginning of the experiment.

Niclosamide Administration.

Niclosamide is dissolved in water and a single dose administered at 3mg/kg to lightly anesthetized mice through a 3.5 Fr catheter insertedinto the rectum.

Pharmacokinetic Study Design.

At 0.25, 0.5, 1, 2, 4, 8 16 hours after niclosamide administration,plasma and colon specimens are collected from 3 mice per time point andhigh-performance liquid chromatography is used to measure the tissueconcentrations niclosamide and its metabolites.

Calculation of Plasma and Colon Ratio.

The mean concentration of niclosamide colon concentration (mg/mg) andniclosamide plasma concentration (mg/mL) is plotted and the ratio iscalculated.

Results.

Niclosamide exhibits a colon to plasma exposure ratio that exceeds 10:1.

Example 6: Doses of Niclosamide that are Therapeutic Against ColitisResult in Colonic Exposure Levels in Mice that are Associated withMitochondrial Uncoupling

Objective.

To determine if niclosamide colonic exposure is associated withniclosamide concentrations that induce mitochondrial uncoupling.

Model.

Results from Examples 2 and 5 are used together. Example 2 is used todefine a dose-response relationship between Niclosamide concentrationand mitochondrial uncoupling. Using the pharmacokinetic data fromExample 5, the maximum Niclosamide concentration in colon is determined.This concentration is directly compared to the dose-response data todetermine if efficacious exposure is sufficient to induce mitochondrialuncoupling in colon.

Niclosamide Administration.

Niclosamide is dissolved in water and a single dose is administered at 1or 3 mg/kg to lightly anesthetized mice through a 3.5 Fr catheterinserted into the rectum.

Pharmacokinetic Study Design.

At 0.25, 0.5, 1, 2, 4, 8, and 16 hours after niclosamide administration,colon specimens are collected from 3 mice per time point per group andhigh-pressure liquid chromatography is used to measure the tissueconcentrations of niclosamide and its metabolites.

Calculation of Target Coverage.

The average area under the colon concentration versus time curve (AUC)and the peak concentration is calculated and is plotted and is graphed.Y axis represents niclosamide concentration in μM while X axisrepresents time. To estimate target coverage, the graph includes ahorizontal line at the level in which niclosamide induces mitochondrialuncoupling in lamina propria T cell.

Results.

At doses below maximal efficacy (e.g. 1 mg/kg), niclosamide does notreach 5 μM concentration in the colon. At therapeutic dosage of 3 mg/kgNiclosamide reaches a colon peak concentration >5 μM. Since 5 μM is theniclosamide concentration able to induce mitochondrial uncoupling morethan 50% of in lamina propria T cell, this data indicate thatefficacious exposures result in colon concentration of niclosamide thatare associated with mitochondrial uncoupling and demonstrate that thetherapeutic mechanism is associated with mitochondrial uncoupling.

FIG. 1. Niclosamide induces cell death in lamina propria T cell fromactive IBD. LPMC (lamina propria mononuclear cells) from IBD subjectswere isolated from macroscopically inflamed intestinal area and treatedwith DMSO or niclosamide (10 μM) for 16 hours. Cell death in laminapropria T cell (CD3+) was determined by measuring 7-AAD staining by flowcytometry.

FIG. 2 includes graphs and images showing that niclosamide exhibitsrobust efficacy in murine TNBS model of ulcerative colitis whenadministered rectally (locally), but not by intraperitoneal injection(systemically).

Example 7: Synthesis of Co-Crystals

A) L-Proline (35.2 mg) and niclosamide (100 mg) are combined in a steelvessel containing a steel ball. To this mixture is added 5 drops ofethanol. The sample is milled for 15 minutes after which time conversionto co-crystal is substantially complete.

The above example is meant to illustrate but not limit the invention.Other methods for achieving the described invention include grindingwith a mortar and pestle, co-milling, slurry conversion, andconcentration of a solution of both components.

It is understood by those skilled in the art that a similar co-crystalcan be produced from D-proline and from mixtures of L- and D-prolinesuch as a racemic mixture thereof.

B) L-Proline (35.2 mg) and niclosamide (100 mg) are combined in a steelvessel containing a steel ball. To this mixture is added 5 drops ofpropylene glycol. The sample is milled for 15 minutes after which timeconversion to cocrystal is substantially complete.

C) Imidazole (20.8 mg) and niclosamide (100 mg) are combined in a steelvessel containing a steel ball. To this mixture is added 5 drops ofethanol. The sample is milled for 15 minutes after which time conversionto co-crystal is substantially complete.

The above example is meant to illustrate but not limit the invention.Other methods for achieving the described invention include grindingwith a mortar and pestle, co-milling, slurry conversion, andconcentration of a solution of both components.

Example 8: Preparation of Enema Formulation Components

The liquid carrier shown in Table 11 below were prepared according tothe following procedure. propyl 4-hydroxybenzoate and methyl4-hydroxybenzoate were dissolved in hot water. The solution was allowedto cool to room temperature, and additional water was added tocompensate water loss due to evaporation that occurred in the priorstep. The sodium salts were added and dissolved under stirring for 10minutes (pH: 6.5-7.5). Methylcellulose and povidone were dispersed usinga turbomixer (9000 rpm, 30′). The preparation was allowed to stand forseveral hours to let foam decant. Typically, the preparation of theliquid carrier was not stored and used immediately. However, whenstored, the liquid carriers were stored in 500 mL polyethylene bottles.The liquid carrier exhibited the properties shown in Table 11.

TABLE 11 Components Quantity (%) Methyl cellulose (Methocel A15C 1.40premium) Povidone (Kollidon K30) 1.00 Propyl parahydroxybenzoate 0.02Methyl parahydroxybenzoate 0.20 Disodium phosphate dodecahydrate 0.15Sodium dihydrogen phosphate dihydrate 0.05 Water purified Up to 100Technological characterization (as IPC) Clear to opalescent Appearancecolloidal dispersion Dynamic viscosity * 41 mPas s pH  7.023 Density1.0075 g/mL

The wet granulation preparations shown in Table 12 were preparedaccording to the following procedure. The internal phase ingredients arecombined and mixed in a high-shear granulator. A granulating solutionwas prepared from water and the indicated agents. This solution is addedto the mixture of the inner phase resulting in the formation ofgranules. Once the granulation was formed and dried, the external phaseingredients were added to the dry granulation. The resultant wetgranulation preparations can be suspended in the above-described liquidcarriers using conventional procedures.

TABLE 12 Niclosamide Strength 450 mg 450 mg Component (%) Inner phaseNiclosamide 100 98.5  77  66  50  61.64 Colloidal silicon — 1.0 —dioxide (Aerosil 200) Magnesium stearate — 0.5 — Cellulose — — 23  34 50  — microcrystalline (Avicel PH101) Crospovidone — — — — — 1.92(Kollidon CL) Lactose monohydrate — — — — — 30.82 (Pharmatose 200M)Granulating Povidone — — — — — 2.74 solution (Kollidon K30) Sodiumlauryl sulfate — — — — — 0.68 Purified water — — — — — * External Talc —— — — — 1.92 phase Magnesium stearate — — — — — 0.27 Theoretical weight(mg) 450 456.9  593.4 692.3 913.8 730.0 *quantity used: 123 mg/units,removed during the process Process Parameter 1) Calibration step rawmaterials Manual calibration 1.1) Calibration sieve Size 1.0 mm 2)Mixing step Turbula, glass container — 2.1) Mixing time - rotation speed5′-34 rmp — 3) Granulation step — Manual granulation 3.1) wet granulatesieve 1.0 mm Technological Characterization Granulate Loss on drying(105° C. for 10′) — 1.4% Final mix Flowability* 10.0 It did not pass Itdid not pass Flow throw an orifice of 15.0 It did not pass 6.1 g/secØ(mm): 25.0 It did not pass 17.8 g/sec Suspendability Not homogeneoussuspendability and Rapid and very poor mixture wettability HomogeneouspH 6.9 *100 g of granulate have to pass through an orifice of increasingsize 10 or 15 or 25 (etc.) mm diameter and the size of the orifice isincreased if the powder is not passing through. When it passes the timeis taken so that the smaller the diameter of the orifice and higher theamount/second the better it is for the flow properties of the granulate.Analytical test Niclosamide assay (%) — 58.84%

Example 9. Niclosamide Suspension Administered Rectally as an Enema hasEfficacy in a Mouse Model of Ulcerative Colitis

Objective:

The objective of this experiment was to determine if niclosamidesuspension administered rectally as an enema to mice with colitisreduces disease activity.

Model:

Intra-rectal administration of trinitrobenzene sulfonic acid (TNBS) tomice results in colitis. TNBS elicits cell-mediated immune responses andinduces transmural inflammation in the gut with morphological andhistopathological features similar to those of human inflammatory boweldisease. TNBS induces diffuse colonic inflammation, characterized byincreased leukocyte infiltration, edema, and ulceration. It is very wellreported that administration of TNBS is associated with predominantactivation of Th1-mediated immune response manifested by increasedcytokines such as interferon-γ (IFN-γ), tumor necrosis factor-α (TNF-α)and interleukin-17A (IL-17A) as well as dense infiltration of CD4+ Tcells. Disease activity in the TNBS model can be determined by loss ofbody weight, histopathological evaluation of the colon showing evidenceof inflammatory damage and evidence of pro-inflammatory cytokinesdetected in colon tissue.

Animals and Treatments:

Studies of TNBS colitis were performed in 8- to 12-week-old male Balb/cmice (Jackson Laboratories, stock number 000651). For induction ofcolitis, 2.5 mg of TNBS (Sigma-Aldrich, Milan, Italy) in 50% ethanol wasadministered to lightly anesthetized mice through a 3.5 F catheterinserted into the rectum. The catheter tip was inserted 4 cm proximal tothe anal verge, and 150 μL of fluid was slowly instilled into the colon,after which the mouse was held in a vertical position for 30 seconds.Mice were exposed to TNBS or 50% ethanol vehicle (EtOH) on Day 0. TNBSor EtOH exposed mice were subsequently dosed rectally with eithernothing, vehicle used for niclosamide (phosphate buffered saline) orniclosamide enema suspension (0.03; 3; 30 mg/kg as indicated) byadministering a 150 μl volume of niclosamide suspension prepared as a 4;0.4; 0.04 mg/ml suspension of niclosamide (Sigma-Aldrich) in phosphatebuffered saline. Niclosamide or vehicle only were administered on day 1and day 2. Weight changes were recorded daily and tissues were collectedfor histologic study and RNA analysis at the end of the study.

Histopathology:

For histologic analysis, tissues were fixed in 10% neutral bufferedformalin solution, embedded in paraffin, cut into tissue sections, andstained with hemaotoxylin & eosin (H&E). For TNBS-induced colitis,stained sections were examined for evidence of colitis and assigned acolitis score (0-5) by considering the presence of acute and chronicinflammatory infiltrates, elongation and/or distortion of crypts, frankulceration, and thickening of the bowel wall.

RNA Extraction, cDNA Preparation, and Real-Time PCR for CytokineDetection:

RNA was extracted from fresh mucosal samples of treated mice usingTrizol reagent according to the manufacturer's instructions (Invitrogen,Carlsbad, Calif.). A constant amount of RNA (1 mg per sample) wasreverse-transcribed into cDNA, and this was amplified using asybergreen-based PCR (Bio-Rad, Hercules, Calif.) using PCR conditionsand primer sequences appropriate for specific detection of IL-17A, IFN-γand TNF-α. β-actin was used as a housekeeping gene to determine relativeexpression. Gene expression was calculated using the ΔΔCt algorithm.

Results and Conclusions—

As shown in FIG. 4A, niclosamide suspension administered rectally at adose of 30 mg/kg on days 1 and 2 results in recovery of body weightinitially last due to TNBS-induced colitis. There is no recovery ofweight in untreated or vehicle control treated mice.

As shown in FIG. 4B, niclosamide suspension administered rectally at adose of 30 mg/kg on days 1 and 2 results in a significantly lowercolitis score compared to vehicle control treated mice or mice thatreceived TNBS and no other treatment, based on H&E analysis of colonbiopsies.

FIG. 4C demonstrates expression of inflammatory cytokines in intestinalbiopsied tissue detected by real-time PCR. TNBS exposure in presence ofvehicle increases expression of TNFa, IFNy and IL-17A compared to EtOHcontrol animals that receive no TNBS. Niclosamide administered rectallyat 0.03, 3.0 and 30 mg per kg body weight dose-dependently reduces thelevel of RNA of each cytokine relative to expression of RNA for β-actin,used as a housekeeping gene for normalization.

The results support the conclusions that rectally administeredniclosamide suspension treats colitis in a mouse model of humaninflammatory bowel disease that recapitulates features of human diseaseincluding colon infiltration by T cells and increased expression ofpro-inflammatory cytokines. The treatment response to niclosamidesuspension administered rectally includes dose-dependent modulation ofpro-inflammatory cytokine gene expression. Collectively, these resultsexemplify the claim that rectal administration of niclosamide suspensionis a treatment for inflammatory diseases of the colon.

Example 10. Niclosamide Reduces the Pro-Inflammatory Potential of TCells Isolated from the Lamina Propria of Human Intestine

Objective:

The objective of this experiment was to determine if niclosamidedirectly reduces the proinflammatory potential of human T cells isolatedfrom the lamina propria sampled as a biopsy from a person withulcerative colitis (UC).

Model:

Lamina propria mononuclear cells (LPMC) in the human intestine arecomprised in part by T cells which mediate pathological processesincluding inflammatory bowel disease. LPMCs can be isolated from humanintestine tissue biopsies. After isolation LPMCs T cells remain viableex vivo under appropriate culture conditions for periods of time thatallow ex vivo experiments. These cells can be used to investigate iftest agents affect their production of pro-inflammatory cytokinesincluding interferon-gamma (IFN), tumor necrosis factor-alpha (TNF) andinterleukin 17A (IL-17A), to determine if a test agent affects thepro-inflammatory cytokines that mediate inflammatory bowel disease,including UC.

Cell Isolation and Culture:

Cells were obtained from colon biopsy specimens of a human from areaswith moderate to severe UC. For the isolation of lamina propriamononuclear cells (LPMCs), the specimens were initially washed in Hank'sbalanced salt solution (HBSS) then cut into 0.5-cm pieces, and incubatedwith stirring in pre-warmed HBSS containing 1 mM DTT at 37° C. for 15minutes. The supernatant was removed and the sample washed with stirringwith HBSS for 5 minutes twice. Samples were incubated with stirring inpre-warmed HBSS containing 5 mM EDTA for 30 minutes. The supernatant wasremoved and the sample washed with stirring with HBSS for 5 minutesthree times. The tissue was then digested further in RPMI 1640containing 2 mg/ml Liberase and 0.01 ug/ml DNase I for 1 hours at 37° C.with stirring. After digestion, the mononuclear cells in suspension werecollected and centrifuged at 400 g for 10 minutes. After two washings inHBS, the pellet was resuspended in a 40% Percoll solution and layered onthe top of a Percoll solution (100%, 60%, 40%, and 30% Percoll inHBSS.). The tube was centrifuged at 400 g for 25 minutes, and LPMCs atthe 60%-40% Percoll layer interface were collected. The isolated cellswere counted and checked for viability using 0.1% trypan blue (viabilityranged from 86% to 94%). Cells were washed out of Percoll with HBSS andresuspended in RPMI 1640 supplemented with 10% heat inactivated FBS, 1%L-glutamine, 100 U/mL penicillin, and 100 mg/mL streptomycin at aconcentration of 1×10⁶ cells/mL and plated in 96-well culture plates(200000 cells/well) (Nat Protoc. 2007; 2(10):2307-11)

Treatment with Test Material Niclosamide—

Niclosamide (purchased from Sigma) was dissolved in dimethyl sulfoxide(DMSO) and added to the culture medium to achieve a concentration of 5μM. Samples were incubated for 24 h at 37° C. A vehicle only control (inplace of niclosamide) was run concurrently.

Measuring Pro-Inflammatory Cytokines.

After treatment with either niclosamide or vehicle control as describedabove, LPMC were stimulated with PMA (10 ng/mL), ionomycin (1 μg/mL),and brefeldinA (10 μg/mL; eBioscience, San Diego, Calif.). After 5 h,cells were stained with the following Abs: anti-CD3-PerCP (1:50, finaldilution, BD Biosciences, San Jose, Calif.) and fixed with 1%formaldehyde for 20′. Subsequently cells were permeabilized with 0.5%saponin in 1% BSA FACS buffer and stained with the following Abs:anti-IFN-γ-PE (1:50, final dilution; clone XMG1.2, BD Biosciences),anti-IL-17A-APC (1:50, final dilution, clone eBio17B7 AffymetrixeBioscience), Anti-TNF-PEcy7 (1:50 final dilution, clone MP6-XT22Affymetrix). Appropriate isotype-matched controls from BD Bioscienceswere included in all of the experiments. A flow cytometer FACSVerse flowcytometer and FACSSuite software [BD Biosciences] was used for toanalyze results.

Results and Conclusion—

Niclosamide at 5 μM causes a decrease in human LPMCs T cells thatproduce pro-inflammatory cytokines including TNF, IFN, and IL-17Arelative to vehicle only negative control (FIG. 5).

Example 11. Administration of Niclosamide Using a Formulation thatResults in a Concentration of Niclosamide in the Rectal Mucosa that isBoth Detectable and Significantly Greater than the Corresponding PlasmaNiclosamide Concentration

Rabbits (New Zealand White KBL Rabbit (SPF: Specific Pathogen Free),naïve to any experimental procedures, Charles River Laboratories S.p.A.Italia.—only males will be used) were treated with a single dose ofniclosamide suspensions containing magnesium stearate and colloidalsilica (98.5% Niclosamide, 1% Silica, colloidal hydrated, and 0.5%Magnesium stearate—manually crushed with mortar and pestle then sievedthrough 60 mesh (250 um) and then suspended in the liquid carrierdescribed in Example 8) at the dose levels specified. Following dosing,blood samples and rectal mucosa was obtained at indicated time points.See Tables 13 and 14.

TABLE 13 Niclosamide Plasma Concentrations (ng/mL) Treatment B 7.5(Study Niclosamide: Evaluation of the Pharmacokinetics following aSingle Rectal Administration to NZW Rabbits) Time Subject SubjectSubject Subject Subject Mean Hours 666 667 668 669 670 ng/ml Std dev 0BLQ BLQ BLQ BLQ BLQ N/A N/A 1 5.79 2.98 2.81 4.13 BLQ 3.9275 1.3731321 25.19 3.26 50.7  19.716667 26.849701 4 BLQ 21.7 2.21 11.955 13.781511 84.73 4.63 BLQ 4.68 0.0707107 24 BLQ 1.07 BLQ 1.07 N/A

TABLE 14 Niclosamide Rectal Concentration (ng/ml) after 1 hour Subejct669 Subject 670 Mean Std dev 12.3 32.8 22.55 14.4957

Rectal administration of niclosamide (7.5 mg) results in mean rectalniclosamide concentration of 22.55 ng/ml (stdev 14.49) compared to aplasma concentration of 3.93 ng/ml (stdev 1.37) 1 hour following dosing.This difference means that the rectal concentration of niclosamide ismore than 5-times the plasma concentration at 1 hr.

Example 12. Niclosamide Reduces Mitochondrial Membrane Potential in TCells Isolated from the Lamina Propria of Human Intestine

Objective—

The objective of this experiment was to determine if niclosamide candirectly reduce the mitochondrial transmembrane potential in T cellsisolated from human intestine lamina propria.

The Model—

Lamina propria mononuclear cells (LPMC) in the human intestine arecomprised in part by T cells which mediate physiological andpathological processes including inflammatory bowel disease. LPMCs canbe isolated from human tissue biopsies. After isolation LPMCs T cellsremain viable ex vivo under appropriate culture conditions for periodsof time that allow ex vivo experiments. These cells can be used toinvestigate mechanisms that regulate their mitochondrial function andsurvival. They contain respiring mitochondria and as such their responseto test agents may be assessed. Uncoupling is identified and quantifiedby a detecting a drop in the electrochemical gradient across themitochondrial inner membrane (ΔΨm).

Cell Isolation and Culture—

Cells were obtained from biopsy specimens of the small or largeintestine or rectum of humans from areas of gastrointestinal tissue withmoderate to severe Crohn's disease (CD). For the isolation of laminapropria mononuclear cells (LPMCs), the specimens were initially washedin Hank's balanced salt solution (HBSS) then cut into 0.5-cm pieces, andincubated with stirring in pre-warmed HBSS containing 1 mM DTT at 37° C.for 15 minutes. The supernatant was removed and the sample washed withstirring with HBSS for 5 minutes twice. Samples were incubated withstirring in pre-warmed HBSS containing 5 mM EDTA for 30 minutes. Thesupernatant was removed and the sample washed with stirring with HBSSfor 5 minutes three times. The tissue was then digested further in RPMI1640 containing 2 mg/ml Liberase and 0.01 ug/ml DNase I for 1 hours at37° C. with stirring. After digestion, the mononuclear cells insuspension were collected and centrifuged at 400 g for 10 minutes. Aftertwo washings in HBS, the pellet was resuspended in a 40% Percollsolution and layered on the top of a Percoll solution (100%, 60%, 40%,and 30% Percoll in HBSS.). The tube was centrifuged at 400 g for 25minutes, and LPMCs at the 60%-40% Percoll layer interface werecollected. The isolated cells were counted and checked for viabilityusing 0.1% trypan blue (viability ranged from 86% to 94%). Cells werewashed out of Percoll with HBSS and resuspended in RPMI 1640supplemented with 10% heat inactivated FBS, 1% L-glutamine, 100 U/mLpenicillin, and 100 mg/mL streptomycin at a concentration of 1×10⁶cells/mL and plated in 96-well culture plates (200000 cells/well) (NatProtoc. 2007; 2(10):2307-11)

Treatment with Test Material Niclosamide—

Niclosamide (purchased from Sigma) was dissolved in dimethyl sulfoxide(DMSO) and added to the culture medium to achieve a concentration of 5μM. Samples were incubated for 60 min at 37° C. JC-1 was purchased fromThermo Fisher Scientific, dissolved in DMSO then added to the test wellsto achieve a final concentration of 10 μg/ml and allowed to incubate at37° C. for an additional 30 min. A vehicle only control (in place ofniclosamide) was run concurrently. A flow cytometer FACSVerse flowcytometer and FACSSuite software [BD Biosciences] was used forquantification of JC-1 fluorescence in CD45+CD3+ cells.

Measuring mitochondrial membrane potential changes (ΔΨm).

JC-1 is a widely used indicator of mitochondrial membrane potential.JC-1 has advantages over other cationic dyes in that it exhibitspotential-dependent accumulation in mitochondria indicated by afluorescence emission shift from green (˜525 nm) to red (˜590 nm).Consequently, mitochondrial depolarization is indicated by a decrease inthe red/green fluorescence intensity ratio. The potential-sensitivecolor shift is due to concentration dependent formation of redfluorescent J-aggregates.

Measurement of and Calculation of Change of ΔΨm in T Cells—

In order to specifically distinguish T cells from other cells in LPMCs,LPMC were stained with anti-CD45 and anti-CD3 antibodies. Anti-CD45monoclonal antibody labeled with PerCP-Cyanine5.5 (Ex488 Em695) waspurchased from Ebioscience (clone 2D1); anti-CD3 monoclonal antibodylabeled with eFluor® 450 (Ex405 Em455) was purchased from Ebioscience(clone OKT3). The anti-CD45 antibody binds human CD45 antigen, that isexpressed in by all hematopoietic cells excluding circulatingerythrocytes and platelets. The anti-CD3 antibody specifically bindshuman CD3 antigen that is selectively expressed on T cells. LPMCCD45+CD3+ T cells were first defined by their fluorescence emission fromlabeling with eFluor® 450-anti-CD3 antibody andPerCP-Cyanine5.5-anti-CD3 antibody. The fluorescence intensity of JC-1detected at ˜525 nm and ˜590 nm in the CD45+CD3+ T cell population wasthen measured.

Results and Conclusion—

Niclosamide at 5 μM causes a decrease in ΔΨm in human LPMCs T cellsrelative to negative control (see FIG. 6).

A number of embodiments of the invention have been described.Nevertheless, it will be understood that various modifications may bemade without departing from the spirit and scope of the invention.Accordingly, other embodiments are within the scope of the followingclaims.

What is claimed is:
 1. A method for treating iatrogenic autoimmunecolitis in a subject in need thereof, the method comprisingadministering an effective amount of a niclosamide wherein theiatrogenic autoimmune colitis is induced by an immune checkpointinhibitor, wherein the immune checkpoint inhibitor targets CTLA-4. 2.The method of claim 1, wherein the immune checkpoint inhibitor is anantibody.
 3. The method of claim 2, wherein the antibody is ipilimumabor tremelimumab.
 4. The method of claim 2, wherein the antibody isipilimumab.
 5. The method of claim 1, the method comprisingadministering an effective amount of niclosamide to the GI tract of thesubject.
 6. The method of claim 1, the method comprising, wherein themethod comprises locally administering an effective amount ofniclosamide to the subject to the GI tract of the subject.
 7. The methodof claim 1, wherein the method comprises topically administering aneffective amount of niclosamide to the subject to the GI tract of thesubject.
 8. The method of claim 1, wherein the niclosamide isadministered by rectal administration.
 9. The method of claim 8, whereinthe niclosamide is administered by enema, rectal gel, rectal foam,rectal aerosol, or suppository.
 10. The method of claim 8, wherein theniclosamide is administered by enema.
 11. The method of claim 1, whereinthe niclosamide is administered by oral administration.
 12. The methodof claim 11 is administered by tablet or pill.
 13. The method of claim1, wherein upon administration, the concentration of the niclosamide inthe GI tract is higher than the concentration of the mitochondrialuncoupling agent in the plasma compartment.
 14. The method of claim 1,wherein the niclosamide in the plasma compartment is subject to firstpass metabolism.
 15. The method of claim 6, wherein upon administration,the concentration of the niclosamide in the GI tract is about 5 timeshigher than the concentration of the niclosamide in the plasmacompartment.
 16. The method of claim 1, wherein upon administration, theconcentration of the niclosamide in the colon is higher than theconcentration of the niclosamide in the plasma compartment.
 17. Themethod of claim 1, wherein upon administration, the concentration of theniclosamide in the colon is about 5 times higher than the concentrationof the niclosamide in the plasma compartment.
 18. The method of claim 1,wherein upon administration, the concentration of the niclosamide in therectal mucosa is higher than the concentration of the niclosamide in theplasma compartment.
 19. The method of claim 1, wherein uponadministration, the concentration of the niclosamide in the rectalmucosa is about 5 times higher than the concentration of the niclosamidein the plasma compartment.
 20. The method of claim 1, wherein thesubject is a human.